Microscopic Exam Body Fluids

CLINICAL MICROSCOPY: Microscopic and Body Fluids MLS INTERNS 2019 NOTES TEAM
THE STEPS FOR PERFORMING A MICROSCOPIC EXAMINATION THAT MEETS FACTS THAT RELATE TO SOURCES OF ERROR IN THE EXAMINATION OF
QUALITY CONTROL STANDARDS URINE SEDIMENT
1. Use fresh urine. If unable to perform within one hour of voiding, 1. Using dirty slides makes accurate differentiation of urine sediment
refrigerate for up to three hours. difficult.
2. Centrifuge a standard quantity. Use either 10, 12, or 15 mL. 2. Refrigerating urine will cause the urine specimen to become turbid
3. Use conical centrifuge tubes. which will make the microscopic examination more difficult.
4. Centrifuge using the same centrifuge, rotor, centrifuge speed and Note: Up to 10% of urine are turbid or cloudy at the time of voiding.
force and time. 3. If dilute acetic acid is used to dissolve amorphous phosphates, then
5. Decant urine the same way or same technique, leaving a residual of any erythrocytes in the urine specimen will be hemolyzed.
0.5 to 1.0 mL urine to resuspend the sediment and always be 4. There are a number of lookalikes that may be found in urine. Oil
consistent in the amount of residual urine remaining. droplets, yeast, certain crystals, and even bubbles have been confused
6. If sediment is to be stained, always add the stain to the sediment in with erythrocytes by inexperienced laboratorians.
the tube, not in the slide. 5. Using a sediment stain will facilitate recognition of formed elements
7. Transfer the sediment to the slide the same way, using the same type in the urine.
of transfer pipette to deliver the same size drop. Do NOT invert the 6. Urine that has stood at room temperature for two or more hours will
tube to transfer sediment. not be reliable as some of the constituents in the urine will have
8. If using a cover glass, use the same size cover glass since a larger or disintegrated or disappeared.
smaller size will affect the way sediment distributes under the cover 7. If urine contains myoglobin, it will yield a positive blood test and no
glass. erythrocytes will be seen.
9. Be consistent in the way that the microscopic evaluation is performed. 8. If a patient takes mega-doses of ascorbic acid, then there may be a
10. Examine the sediment under low power first to detect the presence suppression effect upon the blood, nitrate, glucose, and bilirubin
or absence of casts. reagent test causing a false-negative test result.
11. Use the 40X objective to identify sediment.
CORRECT PROCEDURE FOR REPORTING URINARY SEDIMENT AND/OR
12. Examine the same number of objective fields. Ten fields are
WHEN GIVEN DATA, CORRECTLY COMPELTE A URINALYSIS REPORT
acceptable, but 20 is preferable.
13. Use the same terminology for reporting out sediment. Report should be filled out according to the following parameters.
14. Check microscopic findings with physical and chemical results to
assure accuracy of report. 1.
Renal Tubular Epithelial cells

Transitional Epithelial cells
Crystals
Parasites
USA INTERNS AY 2018-2019 1 of 61

2. EXPECTED STAINED COLOR WHEN THE FOLLOWING FORMED ELEMENTS

Casts ARE STAINED WITH STERNHEIMER-MALBIN (S-M) STAIN
Squamous Epithelial cells
Mucus: Pale pink or pale blue
3. + (trace), 1+ (few), 2+ (moderate), 3+
(many), 4+ (too numerous to count) Erythrocytes
Bacteria Neutral pH pink to purple
Yeast Alkaline pH purple
Mucus Acidic pH do not stain
Fat droplets RBC may not stain well with SM stain
Spermatozoa Leukocytes
Amorphous crystals Nucleus Purple
4. Report casts, crystals, and epithelial cells as to their type. Cytoplasm Purple granules
5. Artifacts, as a rule, are not reported. If you see undigested animal or Bacteria
plant fibers, this should be investigated to rule out if the urine is Motile (alive) not stained
actually stool water, poor hygiene habits, or a vesicosigmoid fistula. Non-motile (dead) purple
Renal Tubular Epithelial cells
Nucleus dark purple
LIST OF ARTIFACTS AND STATE HOW THEY APPEAR IN THE URINE Cytoplasm light shade of purple
Plant cells: via fecal contamination or vesicosigmoid fistula Transitional cells
Starch: body powder from patient or laboratorian gloves Nucleus dark purple
Talc: Body powder from patient Cytoplasm light shade of purple
Pollen: Air contamination Squamous Epithelial cells
Muscle fibers: via fecal contamination or vesicosigmoid fistula Nucleus purple
Cotton fibers: Cytoplasm light shade of purple
collection Trichomonas vaginalis, when alive, has a light greenish
Synthetic fibers: ng urine collection appearance with SM stain
Hair: Hyaline cast: Pale pink or pale blue matrix, similar to mucus
Paper fibers: during urine collection or wiping urine container Waxy cast: Pale pink or pale blue, similar to the hyaline cast
Oil droplets: body lotion, catheter lubricants NOTE:
Wood fibers: Wooden applicator sticks Most casts with inclusions will demonstrate a similar staining
Plastic shards: usually shed off from centrifuge tube reaction in the matrix. The inclusion may or may not stain.
Glass chips: from the coverslip or glass slide due to scratching Color perception varies among techs.
Air bubbles: appear when transferring the specimen to the slide

SUPRAVITAL STAIN: DEMONSTRATE HOW TO PROPERLY STAIN URINE associated with the kidney, there may be an adverse reaction to a
SEDIMENT drug. The presence of increased numbers of eosinophils is a good
indicator that hypersensitive condition is present. Failure to treat
Stain living tissue by perfusing through the cell and distinguishing cell
hypersensitivity may result in renal damage.
components
Provides more detailed images of leukocytes, epithelial cells, casts. PURPOSE OF PRUSSIAN BLUE STAIN
The Sternheimer-Malbin stain and the 0.5% toluidine stain are good
Colors free hemosiderin in urine or that bound in epithelial cells.
stains for urine sediment. SM stain has a tendency to precipitate in
Prussian blue actually binds with the iron molecule in the form of
strongly alkaline urine. If crystals in urine to which SM stain has been
hemosiderin to produce a distinctive blue color
added, such crystals will be brown or purple stellate crystals.
DESCRIBE HOW THE LABORATORY CAN DEMONSTRATE FAT/LIPIDS IN

ERYTHROCYTES IN HYPOTONIC, ISOTONIC, AND HYPERTONIC URINE
URINE
SPECIMENS
The polarizing microscope provides an excellent way to demonstrate
Normal RBC is a round, biconcave disk that appears fractile in urine
lipids.
Cell will have a colorless or yellow green tinge in unstained urine.
It provides greater contrast of urinary sediment and the presence of Diameter will be about 7µm
cholesterol lipids are characterized by the presence of the Maltese If viewed from the side it will take on an hour glass appearance and
cross when viewing from the top, there will be a central area of pallor
Neutral fats do not produce the cross phenomenon ter will be
Lipid stains (Sudan III, Sudan IV, and Oil Red 4) are used by labs to stain absorbed into the RBC (causing it to swell) until it lysis
the neutral fats or triglycerides lipids In isotonic urine, the osmotic forces are equal and the RBC will retain
Polarized light or lipid stains may be used to demonstrate the presence it normal configuration
of free fat globules, oval fat bodies, and fatty casts In hypertonic urine, the osmotic forces will cause fluid to diffuse from
the RBC, leaving the cell to shrink and wrinkle. This cell is called a
USE OF GRAM STAIN TO STAIN URINE SEDIMENT
Gram stain is used in microbiology to differentiate bacteria into gram

positive or gram negative groups. In urinalysis this would also be the
-A- HOW TO DISTINGUISH THEM
purpose.
Yeast cells: Oval and round forms are noted. Variation in size and are
PURPOSE OF
s.
and Oil droplets: Uniform in appearance, but will vary significantly in size.
Used for staining the eosinophils. Eosinophil will appear in the urine in Neutral lipids will stain with lipid stains.
response to hypersensitivity reactions. If the hypersensitivity is
Air Bubbles: Will be variable in size and demonstrate dark ring CLINICAL SIGNIFICANCE OF ERYTHROCYTES IN URINE
phenomenon
Presence if pathological is greater than 5 RBC / HPF
Calcium Oxalate: Monohydrate form of calcium oxalate may contain
oval/round forms. Other (dihydrate) calcium oxalate forms are Normal values are 2 RBC / HPF which is equivalent to < 12 RBC/µL. If
present and easily eliminates this -a- patient is catheterized or experiencing menses
not significant
Pathological implications may be any of the following: (1) bacterial
infection, (2) presence of stones, (3) tumors, (4) trauma, or (5) toxic
with distorted
reactions to drugs or medications
appearances. Their appearance has been reported as bizarre and
consequently their presence may go unreported. TYPES OF LEUKOCYTES AND OTHER PHAGOCYTES THAT MAY BE FOUND IN
Contain variable amounts of haemoglobin which may be randomly URINE
distributed contributing to the appearance of the erythrocyte.
1. Neutrophil: Most common WBC encountered. Cell contains a multi-
Dysmorphic RBC is associated with glomerulonephritis. The RBC will
lobed nucleus, similar in size to a renal tubular epithelial cells and
traverse the length of the nephron, subjected to the osmotic and
ranges in size from 10 to 14 µm. These cells produce the enzyme
physical forces, which produces the dysmorphism. Microscopic look
esterase that cleaves the ester in the reagent strip test pad. Increased
for irregular cell membranes, ring-like forms, blebs, target cells, and
In all inflammatory responses. Report these cells as WBC
other strange configurations.
2. Lymphocytes: Seen in urine specimen and should be reported as
small, mononuclear cells-probably lymphocytes. They do not produce
REPORTED IN THE SEDIMENT esterase and will not affect the reagent strip pad if present in large
numbers. Large numbers are reported in viral infections and acute
Appearance: hazy to cloudy. If <400 RBC/mL, the urine specimen will
glomerulonephritis.
be clear
NOTE: If it is necessary to perform a cytodiagnostic differential,
Color: pink to red to smokey correlation
prepare a smear from the urine sediment an
Sediment button: presence of a red button
Reagent strip test:
urinalysis.
a. Blood pad will be positive
3. Monocytes: Similar in size to Renal Tubular Epithelial cells and are
b. Protein pad may be positive. If bleeding from glomerulus,
easily confused with them. These cells can attain diameters up to 40
albumin is being lost from glomerular capillaries
µm and are easily identified if differentially stained. Report their
presence as large, mononuclear cells-probably monocytes.
4. Eosinophils: Cells are difficult to distinguish from neutrophils. Unless
neutrophils. Eosinophils are increased in acute interstitial nephritis,

allergies, and acute allograph rejection.
5. Macrophages: Also called as histocytes, these cells range usually from COMPARE AND CORRELATE THE URINE MICROSCOPIC FINDINGS OF
30 to 40 µm, but may attain diameters of 100 µm. Nucleus may be LEUKOCYTES TO PHYSICAL AND CHEMICAL PARAMETERS
round, irregular, or kidney shaped. There is an abundance of
m
cytoplasm and vacuoles are present. When seen in urine, they are in
most cases, spherical. Reporting these cells as macrophages. Odor may be strong, pungent or foul
A grey button will appear in the bottom of the centrifuge tube in
NEUTROPHIL AS THE TYPICAL WBC IN URINE pyuria
Clumping will be a characteristic feature in pyuria
Cell should be referred to as a white blood cell, leukocyte or pus cell.
00,000/ m )
In fresh urine specimen, neutrophils will display neutrophil features
and may exhibit amoeboid movement. If nuclear features are Nitrite test = positive (if infective bacteria is a nitrate reducer)
Blood test = positive (if lesions are present due to infection)
on quickly set in and the cell SQUAMOUS EPITHELIAL CELLS IN URINE
becomes increasingly granular in appearance. Nuclei will fuse and the
cell appear as a mononuclear cell. When this occurs, it is easily Most frequently found epithelial cell in urine and has the least clinical
confused with a renal tubular epithelial cells (RTEC). Blebs may appear significance. It is derived from the vagina, prepuce of uncircumcised
on the inner cell membrane, it detach from the cell and floats free. As men, and urethra.
the cells continue to disintegrate, filaments form and extend outward Characterized by an abundance of cytoplasm and a small eccentric
from the membrane surface. When this occurs, the cell membrane nucleus (which is about the size of an RBC). The edges of the cell can
breaks down and the cell ruptures. roll and fold, causing the cell to present in a variety of configurations.
In diluted urine, at room temperature, WBC will swell to form spheres The cytoplasm may appear finely granular and also have a few large
then lysis occurs. Within 2-3 hours, up to 50% of the WBC granules scattered throughout it.
lysed. During the time that the WBC is in its swollen state, cytoplasmic Diameter ranges from 40 to 60 µm. As the cell deteriorates,
granules exhibit Brownian movement and are refractive. When this granulation becomes more prevalent. It is easy to identify and should
occurs, th glitter cell be enumerated with 10x objective. If large numbers of squamous cells
pathological significance. When stained with Sternheimer-Malbin are noted in a urine specimen from a female, this is an indicator of
stain, the glitter cell will appear pale blue. Glitter cells are usually seen vaginal contamination.
in urines with a specific gravity less than 1.019.
Old leukocytes tend to be smaller in size, nuclear features are more When examining the squamous cell, the cell for any aberrant features
indistinct, increased granulation, and in differing states of in the cytoplasm, cell, or nucleus. Comparing the cytoplasm and
disintegration. nuclear ratios will help to identify the squ
cytoplasmic granules, and absence of disintegration

NOTE:

TRANSITIONAL EPITHELIAL CELLS IN URINE CLINICAL SIGNIFICANCE OF TRANSITIONAL CELLS
Also called urothelial or bladder cells, originates from the bladder, Usually there are no clinical significance. In urinary tract infections
urethra of males, ureters, renal pelvis, and the major and minor (UTI) increased numbers are sometimes encountered. If an increased
calyces. number of transitional cells are noted, take time to review the
Size is variable dependent upon which later the cell sloughed from. morphology of cells.
Cells from the outer, superficial layer will be larger and flatter; with a If abnormal morphology is noted, the specimen should be referred to
diameter ranging from 30 to 40 µm, whereas cells from the inner, a pathologist for cytologic review. If the cell originates from the
intermediate layers are smaller and plumper have a diameter of 20 to trigone of the bladder or the renal pelvis, there will often be a tail-like
30 µm. extension.
Transitional cells from the bladder may be larger and closely resemble These cells are called caudate cells and are not diagnostic of anything.
squamous cells.
To recognize the transitional cells:
RENAL TUBULAR EPITHELIAL CELLS (RTEC)
1. Look for larger cytoplasmic granules that tend to accumulate
around the nucleus. This is called nuclear distribution. RTEC is the most significant of the epithelial cells. Originates from any
2. Increased number of inclusions are the rule. part
3. Look for distinct peripheral borders on the cytoplasm and urine sediment as a consequence of natural process of cell
nucleus. replacement. Differentiated by their size and shape. Since it is difficult
4. Shape variations are as follows: to differentiate these cells into proximal, distal, and collecting duct
Round types. To differentiate, slides should be made from the sediment and
Oval
Pear nt only if their numbers are significantly increased.
Caudate Based on the studies of G. B. Schumann, if
Kite per high powerfield, this is a strong indication for renal pathology. The
5. The nucleus will be eccentric, round or oval, and about the size specimen should then be re-evaluated according to lab policy.
of a small WBC. General morphology describes this cell as being round to oblong in
6. It is not abnormal to find cells with two nuclei. shape with a size that is about to two or three times larger than
7. They have the tendency to absorb water and this will alter their leukocytes. Diameters will be as up to 25 µm. A dense, round, often
appearance. eccentric nucleus is typical. The cytoplasm is granular.
8. There will about 6 to 7 times more cytoplasmic area than These cells are not prone to absorb water therefore do not swell and
nuclear area change shape as do transitional cells. They tend to retain their original
shape.

CLINICAL SIGNIFICANCE OF OVAL FAT BODIES CASTS AND HOW THEY ARE FORMED
Oval Fat Bodies (OFB) are renal tubular cells that contain lipids. They Casts are elements of solidified protein that may or may not contain
are formed when the intracellular lipids degenerate and coalesce into inclusions that are found in both normal and abnormal urine. The
lipid globules. If lipids appear in the glomerular filtrate due to distal convoluted tubules and collecting tubules secrete a
glomerular dysfunction and plasma leakage, they will be readily mucoprotein (Tamm-Horsfall Protein) that appears first in the form of
absorbed by the renal tubular cells. fibrils. These fibrils stick to the lumen walls and as more protein fibrils
are secreted, an interweaving occurs and the cast takes forms.
and demonstrate high refractile properties. More easily recognized The formation is augmented if plasma proteins are present, solutes
with the brightfield microscope than the phase microscope. Under low are increased, the pH is aicidc, and filtrate flow through the lumen is
power, they will resemble brownish spheres. Phase microscope slow.
provides a positive identification if the fat globules (with cholesterol Formed in the tubules and conform to the shape and structure of the
and choles tubule. The cast can undergo changes in the tubule and undergo
transitional changes.
must be by a fat stain (Sudan III or Oil Red 4). Have parallel sides but tend to be thicker in the middle and more
The presence of Oval Fat Bodies are pathologically significant and are slender toward the ends. Casts can be long, short, thin, thick,
to be reported out in numbers per high power field. Their presence convoluted, curved, or straight. The cast can be fragile (easily broken)
may suggest any of the following: or resilient (resistant to breaking).
1. Trauma with release of bone marrow fat
HOW pH, SOLUTE CONCENTRATION, URINARY STASIS AND URINE VOLUME
2. Lipid storage
THE NEPHRON CONTRIBUTE TO CAST FORMATION
Niemann-Pick disease),
3. Toxaemia of pregnancy 1. An acidic environment contributes to solute and protein precipitation,
4. Diabetes mellitus which favors cast formation. This occurs most often in the distal and
5. Pyelonephritis collecting tubules.
6. Polycystic kidney disease 2. Solute concentration Salt concentration: favors crystal precipitation
7. Nephritic syndrome and protein precipitation
8. Congestive heart failure 3. Urinary stasis usually occurs for some type of pathological disease
obstruction or congenital abnormality. Stasis facilitates accumulation
When you observe OFB in urine sediment, evaluate the sediment for and concentration of substances that contribute to cast formation.
free floating fat globules. Also look for casts and re-check the protein The presence of plasma proteins (albumin and/or globulins),
test (which should be positive). haemoglobin, or myoglobin enhances cast formation.
NOTE: In lipid storage diseases, fat-filled histocytes and macrophages 4. Oliguria - decrease urine volume
may be observed. They are easily confused with OFB, but are
distinguished by their larger size.
CLASSIFICATION OF CASTS Staining with Sternheimer-Malbin stains enhances their visualization.

They take on pink coloration and borders are more difficult. Numbers
Casts are classified according to t
are increased during severe exercise, dehydration, heat exposure, and
(inclusions) observed in the matrix. Inclusions includes: bacteria, leukocytes,
stress
erythrocytes, renal tubular epithelial cells, lipids, granules, hemosiderin, and
They are also observed to accompany the pathological casts during a
crystals. Casts may be classified as follows:
variety of renal disease, congestive heart failure, and febrile illnesses.
a) Homogenous/non-inclusion: hyaline, waxy Normal values reported by most laboratories are 0-2/LPF. When
b) Inclusion/cellular: leukocytes, erythrocytes, renal tubular epithelial identifying the hyaline cast, do not confuse it with mucus threads.
cells, bacteria
CYLINDEROIDS / CYLINDROIDS
c) Inclusion/non-cellular: lipids, granules, hemosiderin, crystals
d) Pigmented: bilirubin, haemoglobin, myoglobin, drug pigments Cylinderoids are hyaline casts, except that one end (as the rule) will be
tapering or serpentine in appearance. Cylinderoids are to be reported
NOTE: Another category of casts are the broad casts. These casts are extra-
out as hyaline casts.
large and can be classified as identified in all types.
ATHLETIC PSEUDO-
ECOGNIZE THEM
This is a physiological, transient condition that occurs when strenuous
activity is followed by a release of hyaline and granular casts in the
facilitates the precipitation of amorphous phosphates. The precipitate
urine. Once the stress is relieved, the condition disappears.
tends to form in slender, cylindrical appearing arrangements.
To differentiate from true casts, look for the absence of a protein STABILITY AND SOLUBILITY OF CASTS
matrix border: A true cast will present a hyaline-like matrix. Examine
the surround area. If this is a pseudo-cast, there will also be The Tamm-Horsfall protein matrix is stable in an acidic urine with an
indiscriminate masses of precipitate scattered about.
False-casts can also include aggregates of groups of bacteria, cells, or
concentrate the urine and hold an acidic pH, casts may not form. Cast
embedded in a mucoprotein matrix. tend to be fragile, breaking easily. Casts tend to disintegrate in urine
that sits out at room temperature over a period of time.
HYALINE CAST, HOW TO RECOGNIZE IT, AND ITS CLINICAL SIGNIFICANCE
This is the most commonly observed cast and has the least clinical
significance. It consists of a congealed mass of Tamm-Horsfall protein The presence of these casts are always pathological. These casts form
and may contain no or a few inclusions. The refractive index is low and when the RBC becomes trapped in the Tamm-Horsfall Protein matrix
may be easily overlooked if in low numbers. These casts tends to have in a random manner, not in rows or columns. If there is glomerular
rounded ends and present with a variety of sizes and shapes. damage with bleeding, the plasma proteins and fibrinogen will
contribute to the formation of the casts matrix.
These casts form whenever any disorder damages the glomerulus,

nephron, parenchyma tissue of the kidney. These casts are very fragile
The presence of leukocyte casts indicate the presence of a renal
and urine should be fresh and carefully handled to increase the
inflammation or infection.
successs of finding these casts. Since this ca
Specific disorders are
wil quickly deteriorate and the cast becomes unrecognizable as RBC
1. Pyelonephritis
cast. It is then referred to as blood appearance with little color
2. Post streptococcal acute glomerulonephritis
variation.
3. Nephritic syndrome
RBC casts are refractile and have a color ranging from yellow to red-
4. Systemic lupus erythematosus
brown. Look for intact and clearly identifiable erythrocytes. Note the
5. Polyarteritis nodosa
margin of the cast, the edge of the hyaline matrix should be seen
These casts should be reported as number per LPF
it as RBC cast. If RBC casts are present in the sediment, examine the Synonyms are Leukocyte cast or Pus cast
so be present which A positive leukocyte strip test and protein help verify the presence of
helps in the identification of the RBC cast. these casts. If a bacterial infection is present, the nitrate test should
RBC cats are observed in: be positive.
1. Glomerulonephritis Staining the sediment helps to visualize the nuclear detail in the
2. leukocytes in the cast. The consensus among laboratorians is that
3. Polyarthritis casts with leukocytes scattered throughout the cast matrix seldom
4. Sickle cell anemia
5. Sub-acute bacterial endocarditis cast to form a WBC cast by means of it
6. Systemic lupus erythematosus attach to the cast matrix in a random manner and do not form rows or
7. Renal infarction columns. If WBC casts are present, it might be appropriate to
8. After strenuous physical exertion recommend a culture and sensitivity.
RBC casts should be reported as number you see per LPF. A rare RBC CAUTION: Do not confuse WBC casts with Renal Tubular Epithelial Cell
cast may be observed in normal urine. When RBC casts are observed (RTEC) casts.
in urine sediment, it is probable that the blood values for uric acid, GRANULAR CAST AND ITS CLINICAL SIGNIFICANCE
blood urea nitrogen (BUN), creatinine will be increased. If RBC are
present, the strip reagent test pads should be positive for blood and Granular casts are acellular casts and are generally thought to be a
possibly protein. stage in the degeneration process of cellular casts. The size of the
NOTE: RBC and Hemoglobin casts at one time were thought to granules in this cast varies resulting in two general categories: fine
represent different disorders and should be differentiated in the and coarse granular casts. Neither is more significant than the other,
report. This is no longer true and the RBC cast, blood cast, or therefore, the size of granules are not differentiated and these casts
haemoglobin cast may be reported out as RBC cast. are simply reported out as granular casts per lpf.

It is not uncommon to find granular casts accompanying hyalines cast o The urine strongly foams when shaken due to increased
in times of physical exertion, emotional stress, dehydration, or heat albuminuria
stress. It is the second most commonly occurring cast in urine The presence of these casts indicates the following type of renal
sediment. When viewed in the brightfield microscope, these casts pathology:
appear colorless or yellow in appearance. If you observe a hyaline o Nephritic syndrome
cast with several obvious granules, but the cast is predominately o Lipid necrosis
hyaline, then report the cast as the hyaline type. o Diabetic neuropathy
This cast has a high refractive index and is most often observed in the o Lupus neuropathy
cigar shape. This cast will take on a variety of colors dependent upon
the staining material. o Any chronic renal disorder
o Sternheimer-Malbin stain = causes dark blue coloration of o Renal tubular cell death
the granules Report this cast as numbers per LPF
o Bilirubin = will stain the cast a yellow-orange color
WAXY CAST, HOW TO RECOGNIZE IT, AND ITS CLINICAL SIGNIFICANCE
o Phenazopyridium = produces a orange-red color
An acellular cast with a very high refractive index. Its presence in a
FATTY CAST AND ITS CLINICAL SIGNIFICANCE
urine specimen is an indicator of renal tubular damage, severe stasis
This cast may contain fat oval bodies or fat globules or both when and is a very serious pathological finding. These casts are associated
present is pathologically significant. It is characterized by a high with chronic renal failure, nephritic syndrome, diabetic neuropathy,
refractive index, stains with Sudan III or IV (if triglycerides are renal allograft rejection, and renal amyloidosis.
present) and the polarized microscope demonstrates the Maltese In the brightfield microscope, the waxy cast is homogeneously
cross phenomenon (if cholesterol and its esters are present) smooth in appearance, has parallel sides with sharp margins (in
The following may be observed when fatty casts are present: which cracks, fissures, convolutions, and notches can be observed),
o Variable size fat globules within the cast ends that are often blunt and a broken off appearance, and will
o Will NOT take up Sternheimer-Malbin stain appear to have thickened areas.
o Cast matrix will be either hyaline or granular in type The color of the waxy cast in unstained sediment is from colorless to
o Proteinuria is present gray to yellowish. If stained with Sternheimer-Malbin stain, the waxy
o Variable size free floating fat globules in the cast appears pink. Because waxy casts appear in urine specimens
with a serious pathology; these casts often appear with diameter 2 to
o Strongly positive protein test on the reagent test strip or 3% 6 times larger than the average casts.
SSA test These casts are broad casts or renal failure casts.

Note: Performing a Gram Stain may facilitate

recognition of the bacterial cast.
A mixed cast is one that has more than one component. It is not
unusual for more than one component to be incorporated into a PIGMENTED CAST AND ITS CLINICAL SIGNIFICANCE
Tamm-Horsfall matrix. Clinical significance of finding mixed casts will
correlate to that described for a specific constituent. Mixed casts The pigmented cast is usually a hyaline cast that has a distinctive
may be any one of the following components as shown in the coloration form:
following examples: o Bilirubin
o Hemoglobin
o WBC cast + granular cast
o WBC cast + RBC cast o Myoglobin
o Phenazopyridium
Other examples of mixed casts would be:
o Any strongly colored medication
o Fatty + renal tubular epithelial cell
Ordinary urine pigments do not stain casts.
o Bacterial + renal tubular epithelial cell
o Bacterial + RBC If the pigment is due to the haemoglobin, this could be an indicator
o Granular + WBC + renal tubular epithelial cell of a transfusion reaction or haemolytic anemia.
o RBC + renal tubular epithelial cell If the pigment is due to myoglobin, then muscle trauma or acute
renal failure may be the cause
BACTERIAL CAST AND ITS CLINICAL SIGNIFICANCE If bilirubin is present, not only will the casts be stained, but also the
cellular elements. Bilirubin is an indicator of hepatitis.
Bacterial casts may be expected to occur whenever a patient is
Strongly colored medications such as phenazopyridium is an
diagnosed with pyelonephritis. The bacteria are often difficult to
indicator that a treatment process is in progress and may not be
discern within the cast matrix and it is not unusual for a bacterial cast
considered to be clinically significant.
to be reported out as a granular cast.
The following information will help to identify the bacterial cast: SEDIMENT?
o
o Telescoped urine is a term that describes the appearance of all types
in the cast, in which case, the cast is a mixed cast. Because of casts and any other pathological components in the urine
of the responsiveness of neutrophils to bacterial presence, sediment.
few true bacterial casts are observed. It has been used to describe the sediment of patients diagnosed with
o WBC casts may be present. acute glomerulonephritis. Its original meaning described the
o The protein, nitrite and leukocyte esterase pads on the simultaneous appearance of WBCs and RBCs in urine.
reagent strip should be positive. The term now includes the appearance of casts, including broad
casts, waxy casts, and oval fat bodies.

etrical
appearance. They vary in width and length, appearing ribbon-like and
Broad casts occur when the flow of urine in the lumen of the tubules
often wavy. The ends are typically irregular, pointed, or frayed (split).
becomes very compromised. Formation usually occurs within dilated
Mucus can easily overlooked in the urine specimen because of its low
or atrophied distal tubules and the larger collecting tubules. Their
refractive index.
presence is an indicator of a poor prognosis. They may be of any type
Sternheimer-Malbin stain enhances its appearance giving it a pink or
of cast, but granular and waxy types are more often observed.
blue cast. Mucus has been confused with hyaline casts, but careful
CRYSTAL CASTS AND THEIR CLINICAL SIGNIFICANCE observation will reveal the irregular nature of the mucus strands.
Mucus can be seen in clumps.
Crystal casts occur when solutes precipitate in the lumen of the renal
tubules and become trapped in the hyaline cast matrix. It is the YEAST CELLS IN URINE AND ITS CLINICAL SIGNIFICANCE
consensus of most professionals that these casts have no clinical
Yeast are fungi and the most common encountered species in urine
significance.
is Candida albicans. The species is observed most often in diabetic
The two most commonly encountered crystal type casts are calcium
patients and women with vaginal moniliasis. It is possible for a UTI to
oxalates and sulphonamides, with uric acid crystals in third place.
be caused by yeast.
Casts with amorphous urates have been reported. Before reporting
Yeasts are oval to round, small to moderate size, refractile, colorless
out a crystal cast, be sure that you confirm the presence of a protein
cells that resemble erythrocytes. Yeasts are easily differentiated from
matrix and that these are not crystals aligned along a sticky strand of
RBCs because they will not dissolve in dilute acid nor take up vital
mucus. Because crystal casts may cause irritation in the tubules,
stains.
bleeding may occur. Some degree of hematuria may accompany such
Other conditions in which yeast may be found are pregnancy,
casts in urine.
immune-compromised patients, and women on oral contraceptives.
MUCUS IN URINE AND IT CLINICAL SIGNIFICANCE Caution must be exercised in identifying a yeast infection. Yeasts are
present in the air and on the skin. Because of this, yeasts can easily
Mucus is a proteinaceous fibrous material produced and excreted by
contaminate a urine specimen.
the glands in the lower parts of the urinary tract and it also comes
Candida albicans is an oval yeast measuring 2-3 by 4-6 uM. This yeast
from the vagina as a urine contaminant.
ins
Tamm-Horsfall protein is known to be a component of mucus and
gram-positive. It produces budding cells that may be elongated and
renal tubular cells do contribute the presence of this formed
resembles hyphae. Such elongations are called PSEUDOHYPHAE.
element. Mucus is not considered to be clinically significant. There
This organism has the potential to invade the body producing sepsis,
may be clinical concern, if there is an inflammatory condition that is
thrombophlebitis, endocarditis, or other infections of body organs.
characterized by excretion of larger quantities of mucus.

Such invasions are not a problem to the normal and healthy observed in women, usually the result of vaginal contamination.
individual as the immune system readily destroys this yeast.
Individuals with predisposing factors as: o Look for quick, flitting or jerky motions. Although it is
o Diabetes mellitus responsible for vaginal infections, it can also infect the
o General debility urethra, prostate, periurethral glands and bladder.
o Immunodeficiency (AIDS or chemotherapy) o It is a flagellated, pear-shaped protozoan, about the size of
o Indwelling urinary catheters leukocytes or renal tubular epithelial cells. Sizes have been
o Indwelling I.V. catheters reported from 5 uM and up to 30 uM. It has single nucleus,
May find this organism a problem four anterior flagella, an anterior undulating membrane
Two other yeast species can be found in urine, but on an infrequent (ending halfway down the length of the parasite) and a
basis. Torulopsis glabrata are small yeast cells, do not form posterior axostyle.
pseudohyphae, easily phagotized by WBCs. Candida parapsilosis is o It is transmitted sexually producing symptoms in women,
usually found in patients with indwelling catheters. They are but is generally asymptomatic in men. The parasite is fragile
commonly found on the skin and in the feces of healthy patients.
is difficult to identify because it assumes a spherical form
POTASSIUM HYDROXIDE PREPARATION AND ITS ROLE IN URINE TESTING and appears as a degenerating leukocyte or renal tubular
Potassium hydroxide (10% KOH solution) is added to a slide epithelial cell. Staining does not enhance its morphology
preparation of vaginal secretions, slightly diluted with saline. The
slide is warmed and observed macroscopically. KOH enhances fungal be observed in fresh urine.
elements by destroying other cellular and formed elements. Bacteria The other urinary parasite is Schistosoma haematobium, a
are not affected by the KOH solution. The procedure required that a trematode (fluke). It is endemic to the Middle East, Africa and
vaginal swab be placed in 0.5 mL of saline and swirled to mix. A drop Portugal. It is seldom seen in the United States. The fluke resides in
of this combination is placed on a glass slide and a drop of KOH is the venous plexuses of the urinary bladder, prostate gland, uterus
added, mixed, and a coverslip placed over the mixture. The specimen and vagina. Its eggs migrate through the bladder wall and appear in
is then examined under the microscope. the urine. When looking for ova, it is recommended that a late
morning urine specimen be collected and examined.
PARASITES IN URINE AND THEIR CLINICAL SIGNIFICANCE Another parasites occasionally found in urine is the Enterobius
A variety of parasites may be observed in urine. The most commonly vermicularis ovum. If it is present, it is due to fecal contamination. It
encountered parasite is Trichomonas vaginalis. Commonly called a is occasionally seen in urine in the larval form.
most often At one time or another, intestinal parasites have been found in urine.
This would be due to a vesicosigmoid fistula or fecal contamination.
Examples include:
o Entamoeba histolytica gram-stain technique may be helpful in visualizing these

o Ascaris lumbricoides microorganisms. If bacteria are found in a clean catch specimen
o Giardia lamblia collected in a sterile container, a UTI is possible. If bacteria can be
TWO WAYS THAT FECAL CONTAMINATION CAN OCCUR IN URINE possible UTI.
VESICOSIGMOID FISTULA: A pathological occurrence when the To reliably identify bacteria in urine, use a clean-catch specimen
urinary bladder and sigmoid colon fuses and an abnormal opening collected in a sterile container, centrifuge and gram-stain. If
(fistula) forms. The movement of fecal material into the urinary bacteriuria is obvious, but WBCs are absent, investigate the
bladder will then occur. To demonstrate this anomaly may require collection and handling methods to eliminate contamination.
ingesting inert, indicator substance (as charcoal). The presence of
CRYSTAL FORMATION IN URINE
charcoal in the urine is diagnostic.
Improper collection technique is seen most often in infants and Crystals precipitate in urine due to high concentration of solutes.
physically compromised patients. Proper care in cleaning and Factors that causes crystal formation:
collecting the urine will eliminate this problem. o Solute concentration: Factors include dehydration, dietary
excesses, and medications.
BACTERIA IN URINE AND THEIR CLINICAL SIGNIFICANCE o pH: Solubility is pH dependent. Crystals that precipitate in
Normal, healthy urine is sterile and free of bacteria. neutral or alkaline urine are less soluble than the crystals
Most bacteria observed in urine is due to pollution that is a result of that precipitate in acidic urine. As a rule, inorganic salts
careless handling, use of non-sanitized containers, vaginal (calcium, phosphate, ammonium, and magnesium)
contamination, or resultant of gastrointestinal tract contamination. precipitate in alkaline urine. Organic solutes (uric acid,
In the case of contamination, the presence of bacteria is not clinically cystine, bilirubin, and x-ray dye) tend to precipitate out in
significant. acidic urine.
The most commonly encountered bacteria are the gram-negative o The rate of flow through the tubules affects crystal
rods. If bacteria are observed, and there are leukocytes present in formation: A slow rate of flow produces concentrated urine
the sediment, and the chemistry tests for protein, nitrate and and promotes crystal formation. A rapid flow rate produced
leukocyte esterase are positive, then a UTI is possible and a culture more dilute urine and decreased crystal formation.
and sensitivity should be performed. o Temperature: If warm, the solutes remain in solution better
Bacteria observed range for small cocci and rod forms to large forms. and crystallization is retarded or does not occur. If cold,
On occasions large and long bacteria are observed with a large then solutes become less soluble and crystallization occurs
central swelling, these are protoplasts and are the outcome of readily.
antibiotic therapy. If there is difficulty in recognizing bacteria, the

Normal, healthy urine seldom contains crystals. The presence of o If large calcium carbonate crystals are present, they will
take on a dumbbell-like shape. If a dilute acid is added,
the urine at the time of voiding, it is possible that this may be effervescence will be observed as CO2 is given off. Both of
clinically significant. Most crystals observed in urine precipitate out these crystals appear colorless when viewed with the
after sitting (especially if the urine is refrigerated before testing), microscope.
because the solute concentration is high, a super saturated solution,
and the solubility threshold is exceeded as the urine cools. o Macroscopically, these crystals will appear as a pink, brick-
like dust. Because of their chemical nature, they readily
AMORPHOUS SEDIMENT IN URINE AND ITS CLINICAL SIGNIFICANCE
absorb the urinary pigments. It is uroerythrin that imparts
There are three types of amorphous sediments. Each has the the reddish color. In the microscope, they will appear
following details: colorless or sometimes a brownish coloration. These
Precipitated salts crystals are uric acid salts of sodium, potassium,
Not clinically significant magnesium, or calcium.
Are coarse granular in appearance o If these crystals are warmed to body temperature or to
o If the amount of amorphous sediment is abundant, it can 60oC, they will dissolve. They will also dissolve in dilute
make the microscopic evaluation difficult; hence, they alkali. If you add a strong mineral acid like HCl or glacial
literally means without any form . These crystals are period of time, uric acid will crystallize out.
shapeless and formless, resembling sawdust or sand.
LISTS OF NORMAL URINARY CRYSTALS AND THE pH AT WHICH THEY ARE
Amorphous sediment is the most commonly encountered
FOUND:
types of crystals in urine. If urine is tested within the first
hour after collecting (without refrigerated), amorphous Amorphous urates (pH: acid to neutral)
formation is minimized. Calcium oxalate (pH: acid to neutral, sometimes can be observed in a
Neutral to alkaline urine, there are two types of amorphous crystals: slightly alkaline pH)
o Uric acid (pH: acid to neutral, sometimes can be observed in a slightly
observe, urine specimens in lab, a urine with a moderate alkaline pH)
amount (or larger) of amorphous phosphates present, the Monosodium urates (pH: acid to neutral)
macroscopic, cloudy appearance will be white. Calcium oxalate, both di- and monohydrate forms (pH: acid to
o They are soluble in dilute acids and will not dissolve when neutral, sometimes can be observed in slightly alkaline pH)
heated to 60oC. These are made up of magnesium and Amorphous phosphates (pH: neutral to alkaline)
calcium phosphates. Triple phosphate (pH: neutral to alkaline)

Dicalcium phosphate (pH: neutral to alkaline) URIC ACID CRYSTALS AND THEIR SIGNIFICANCE
Calcium phosphate (pH: neutral to alkaline)
The most pleomorphic of the crystals found in urine. Forms/patterns
Calcium carbonate (pH: neutral to alkaline)
include rhombic (diamond), cubes, rosettes (when multiple crystals
Ammonium biurate (pH: neutral to alkaline) cluster and fuse), needles, wedge, dumbbells, hexagons, and
Calcium sulfate (pH: acidic) irregular plates/shapes.
CALCIUM OXALATE CRYSTALS AND TEHIR SIGNIFICANCE IN URINE Uric acid crystals, when first formed are colorless, but because of
their chemical properties, will adsorb pigments from the urine. Uric
Colorless and do not absorb acid crystals will appear in varying shades of yellow or yellow-brown,
pigments from the urine. Most common shape is the octahedral form dependent upon the amount of pigment in the urine. This coloration
which appears as two pyramids joined at their base. When focusing is a key to their identification. Uric acid crystals are quite variable in
on this crystal, there is the appearance of a refractile cross or star in size.
the center of a cube. These crystals are soluble in alkali but are insoluble in acids or
alcohol. Generally, the presence of uric acid crystals are considered
persistent finding of numerous calcium oxalate crystals could be an to be clinically insignificant. In any condition (examples: gout,
indicator of small bowel disease, urinary calculi, renal failure disease, leukemia, lymphoma) in which there is an increase in the cellular
diabetes mellitus, high milk intake, bone fractures, CNS injuries, turnover rate, uric acid crystals will be increased. If a patient is on
ethylene glycol poisoning, or acetazolamide therapy. cytotoxin therapy, there will be an increase in cell destruction. This
These crystals are formed from the calcium salts of oxalic acid and means that purine metabolism will be increased and uric acid crystal
other oxalates. Foods high in oxalic acid and oxalates are: oranges, formation will occur.
cabbage, rhubarb, asparagus, brussel sprouts, tomatoes, spinach,
broccoli, garlic, and berries. SODIUM URATE CRYSTALS AND THEIR CLINICAL SIGNIFICANCE
Two basic types of calcium oxalate crystals: Dihydrate and Sodium urates are a variant of uric acid crystals. Colorless (most
Monohydrate forms. The dehydrate form tends to form squares and often observed) to slightly yellow rods or slender prism. Some
rectangles, whereas the monohydrate form tends to form oval and laboratorians call these uric acid spears. Found singly or in clusters.
dumbbell shapes. Biconcave disk forms have been reported. Dissolved at 60oC. Clinically insignificant and may be reported as
CAUTION: Monohydrate may form long ovals and closely resemble will
acetaminophen crystals. Because there are a variety of forms, these change to the uric acid form.
crystals may be described as being pleomorphic. Regardless of the
type, either form is generally considered to be clinically insignificant.
Calcium oxalate crystals are soluble in dilute hydrochloric acid but
not dilute acetic acid.

TRIPLE PHOSPHATE CRYSTALS AND THEIR CLINICAL SIGNIFICANCE CALCIUM CARBONATE CRYSTALS AND THEIR SIGNIFICANCE
Also called ammonium, magnesium phosphate crystals, Crystals appear most often as an amorphous form. On occasions,
- they will appear in crystalline form and then they are dumbbell in
Often appear as three- or six-sided prism, colorless, and very shape. It has been suggested that this shape is due to clumping and
- fusing of the amorphous crystals. Soluble in dilute acetic acid and will
fern-like forms, but these forms are uncommon. Their presence is effervesce.
generally non-significant, however, they are observed with chronic
AMMONIUM BIURATE CRYSTALS AND THEIR CLINICAL SIGNIFICANCE
UTI, obstructive uropathy, and urinary calculi. These crystals are
characterized by imperfections (not perfectly formed). They are very heir
variable in size and are soluble in 10% acetic acid. shape is peculiar for it fused spheres, tortuous shape, and the
presence or absence of spiny projections. Yellow brown in color,
DICALCIUM PHOSPHATE CRYSTALS AND THEIR SIGNIFICANCE
rarely occurring in fresh urine. If alkaline urine is allowed to stand,
An uncommon variation of calcium phosphate and may be found in then these crystals may precipitate out. They are not clinically
slightly acidic urine. The correct designation is dicalcium hydrogen significant and will dissolve at 60oC, in acetic acid, or sodium
phosphates. They tend to be long slender prisms, with one end hydroxide. Confused with yeast cells and leucin crystals. If
pointed. They are often found in clusters and for this reason may be concentrated HCl is added, these crystals can reform to uric acid
crystals.
and are clinically insignificant.
ABNORMAL CRYSTALS THAT ARE OF METABOLIC ORIGIN
CALCIUM PHOSPHATE CRYSTALS AND THEIR SIGNIFICANCE
Hippuric acid (pH: mostly alkaline, sometimes in acidic and neutral
Crystals are usually observes as large, colorless, irregular, thin plates urine)
that are granular in appearance. They float on top of the urine and Cystine (pH: acidic)
Tyrosine (pH: acidic)
shaped prisms. These are soluble in 10% acetic acid. Leucine (pH: acidic)
CAUTION: Small plates may resemble a degenerate squamous Cholesterol (pH: acidic)
epithelial cell. Bilirubin (pH: acidic)
Hemosiderin (pH: acidic)

HIPPURIC ACID CRYSTALS AND THEIR SIGNIFICANCE

precipitates as crystals. The appearances of these crystals are
Not discussed in most textbooks.
clinically significant and indicate some type of pathology. Cystine
Considered to be an abnormal, but non-pathogenic crystal.
crystals are also observed in pyelonephritis, heavy metal
Formed ad a colorless crystal, and like uric acid, it has an affinity for
acidosis.
urinary pigments to take on a yellowish color.
Occurs most often as an elongated prism with variable lengths and TYROSINE CRYSTALS AND THEIR CLINICAL SIGNIFICANCE
widths, the ends being triangular or pointed.
Tyrosine crystals are fine, delicate needles that may be arranged in
Other shapes reported are needles (which can arrange in clusters),
sheaves or clumps. Sometimes these crystals may have a fine, silky
and rhombic plates.
appearance. They are formed first as colorless crystals and will take
Hippuric acid can be a uric acid look-a-like.
on a pale yellow color if bilirubin (or some other strong dye,
It is soluble in acetic acid (uric acid is not soluble), hot water, alkali,
Sternheimer-Malbin stain will dye the crystals purple) is present.
and ether.
When focusing with a microscope, these crystals may appear black.
It has been reported to be observed in the urine of -
These crystals are soluble in alkali, mineral acids, acetic acid, or heat;
CYSTINE CRYSTALS AND THEIR SIGNIFICANCE but insoluble in aceton, alcohol, or water. Because of their
insolubility in water, these crystals may be seen more often than
A rarely observed crystal that appears in the urine as a hexagonal
plate and has the tendency to laminate to other cystine plates. Sides Water). These crystals are observed with severe liver disease,
of the hexagon may be of unequal lengths and will vary greatly in oasthouse disease, hereditary tyrosinosis, bilirubinuria, or if leucine
size (measuring up to 100 uM) crystals are present.
Crystal is soluble in dilute HCl, ammonia, dilute bases; but is
insoluble in acetic acid, alcohol, boiling water and ether. Cystine can LEUCINE CRYSTALS AND THEIR CLINICAL SIGNIFICANCE
crystallize out in the kidney to form stones, and cause urinary
Often describes as yellow to brown, oily appearing spheres, with
blockage. It is rapidly destroyed by bacteria. When searching for
variable sizes, having the appearance of concentric rings (with or
these crystals, use in fresh urine. Bacteria will rapidly destroy
without radial striations), and a central nidus.
cystine. Avoid letting the slide dry, crystals will wrinkle and change
Their presence is a serious prognosis. These crystals may be observed
in severe liver disease. But more likely to be found associated with
nitroprusside) is used to identify the crystal.
metabolic disorders.
The presence is due to an inborn error of metabolism in which there
Crystals are soluble in hot alkali, hot alcohol, and boiling glacial acetic
is a defect in the transport mechanisms for cystine, ornithine, lysine,
acid. They are insoluble in dilute HCl and warm dilute acetic acid.
and arginine. Cells cannot reabsorb these amino acids and thus
They may be found in urine with tyrosine crystals.
appear in the urine in large quantities and of these four, only cystine
CHOLESTEROL CRYSTALS AND THEIR CLINICAL SIGNIFICANCE HEMOSIDERIN CRYSTALS AND THEIR SIGNIFICANCE
If observed in urine, it is pathologically significant. Cholesterol Hemosiderin, if it precipitates out, will appear as amorphous urates.
crystallized out as broad, flat plates, often characterized by a If hemosiderin crystals are present, they may be confirmed with a
notched corner. These crystals will appear colorless or take on a Prussian blue stain. The presence of hemosiderin in the urine, either
pale-green to yellow coloration. free or crystalline form, is an indicator of an intravascular haemolytic
Soluble in chloroform, ether, and boiling alcohol; but insoluble in episode.
warm alcohol. If cholesterol crystals are truly present, look for a
ABNORMAL CRYSTALS OF IATROGENIC OR DRUG ORIGIN AND INDICATE
specific gravity <1.035 and a positive protein test along with the
THE pH AT WHICH THEY ARE FOUND
presence of fat globules, oval fat bodies, and/or fatty casts. If these
are not present, then another crystal should be considered. Be Penicillin (pH: acid to neutral)
cautious in identifying these crystals since they can resemble Acyclovir (pH: acid to neutral)
radiographic dye crystals. It is recommended that cholesterol Sulfonamides (pH: acid to neutral)
crystals should not be reported if the confirmatory findings are not Radiographic (pH: acid to neutral)
present.
PENICILLIN CRYSTALS AND THEIR CLINICAL SIGNIFICANCE
Cholesterol crystals are observed in lymph gland disorders, chyluria,
severe UTI, and nephritic syndrome. Penicillitype antibiotics are seldom observed in urine. If they should
be present, it will be due to a high dosage when the physician is
BILIRUBIN CRYSTALS AND THEIR SIGNIFICANCE aggressively treating for an infection like meningitis, and
septicaemia. Ampicillin crystals are long, thin, colorless prisms or
Bilirubin crystals have been reported in a variety of forms: fine
needles that may appear singly or in clusters. Penicillin-G crystals
needles (that may form clusters), rhombic plates, cubes, and
tend to be rectangular, oblong and with pointed ends. These types of
granules. Colors range from yellow-brown to reddish-brown.
crystals tend to form when refrigerated .
Soluble in alkali, acetone, chloroform, and acids; but insoluble in
alcohol and ether. Bilirubin is a strong dye and if present in urine, ACYCLOVIR CRYSTALS AND THEIR CLINICAL SIGNIFICANCE
will stain other crystals (especially uric acid) along with cells and Acyclovir is an anti-viral medication. When given high doses of the
casts. The appearance of precipitated bilirubin has the same drug, the urine may demonstrate fine, slender needles that closely
significance as a positive reagent strip test or Ictotest. resemble sodium urate crystals. These crystals are most likely to be
observed in neutral or slightly alkaline urine.
NOTE: When medications are administered in high doses, there is a
chance that crystals may appear in urine. When encountering
unknown crystals in a urine sediment examination, refer to the
patients chart as a strategy to identify the crystals.
SULFONAMIDE CRYSTALS AND THEIR SIGNIFICANCE D. Acetylsulfadiazine

This is a seldom used medication
Its crystals appear as yellow-brown sheaves of
The crystal that is encountered is determined by the drug
wheat with eccentric bindings
e. They precipitate out as colorless
they tended to be insoluble and could precipitate out in kidney,
crystals, but have an affinity to absorb urinary pigments and which
posing a risk for renal damage. Current manufacturing technology
will contribute to a yellow coloration. If a urine is allowed to stand
has essentially eliminated this problem and such crystals are seldom
and the pH becomes alkaline, sulfa crystals tend to dissolve. Fresh
observed in the urine, however a potential risk is still present if the
urine should be used to look for these crystals. To help identify
patient is allowed to become dehydrated. Crystal formation in the
suspected sulfa crystals, review the patients medication charts.
tubules may cause hematuria or oliguria (by blockage).The variety of
settes, sheaves (with RADIOGRAPHIC DYE CRYSTALS AND THEIR CLINICAL SIGNIFICANCE
central or lateral bindings), fans, hexagonal plates, rectangular
These are water soluble radio-opaque chemicals that are readily
plates, arrowheads, rhomboids, and spheres. The following are four
excreted by the kidney. These dyes are derived from triiodobenzoic
acid. If the dyeis designated as a meglumine, it is a triiodobenzoic
A. Sulfadiazine
acid conjugated with a synthetic organic compound to form a more
Appears as spheres with irregular striations, but
water-soluble and less-toxic chemical.
may look like a dense brown spheres or bundles of
needles similar to sheaves of wheat. Triiodobenzoic acid base dyes are a variety of mixtures (with
It has been reported as a look-a-like to the different inorganic anions) and are known by trade names as
ammonium biurate crystal Hypaque, Renograffin, Cystograffin, and Renovist. Crystals of
B. Sulfamethoxazole Renograffin (meglumine diatrizoate) will appear in urine shortly after
injection and may be mistaken for cholesterol crystals.
and is rarely seen. One characteristic of the presence of these radio-opaque chemicals
When observed as a crystal, it appears as a brown in urine is a positive sulfosalicylic acid test. The importance of
sphere that may be unevenly divided or rosettes. recognizing radiographic dye crystals is that of false identification.
C. Sulfasalazine Things to remember about these type of crystals are
A poorly absorbed sulfa that is used to treat enteric 1. They may have appear in urine for up to four hours after
diseases. If there is tissue damage in the intestine, it injection
may be absorbed into the blood stream 2. The strip reagent test for proteinis negative, but 3% SSA test is
It is excreted as sulfapyridine or acetylsulfapyridine likely to be positive
Crystals will appear as rhomboids. 3. Specific gravity will be elevated, usually over 1.040
There is usually no clinical significance associated with these
crystals. If a patient has a kidney disorder or dehydrated, then the

patient may experience a problem. If the crystals are unusually 1. AMNIOTIC FLUID
abundant, the appearance of the urine specimen may be cloudy. Liquid that surrounds the unborn baby(fetus) during pregnancy. It is
contained in the amniotic sac. Commonly called the BAG OF
WHAT IS MEANT WHEN SOMETHING IS OF IATROGENIC ORIGIN?
WATERS, sometimes the membranes, is the sac in which the fetus
Iatrogenic means (a) Physician induced (b) Caused by treatment or develops in amniotes. It is thin but tough transparent pair of
medications. The implication is that an adverse mental, chemical, or membranes that hold a developing embryo (and later fetus) until
physical condition was induced by the effects of treatment. This shortly before birth.
means that the effects could have been avoided by proper care on Formed from the placenta
the part of the physician. Formation in the amnion is regulated by balance between the
production of fetal urine and lung fluids and the absorption from
the swallowing and intramembranous flow.
COMPOSITION
98% water
2% solid substances
Inorganic & organic salts
Fetal epithelium
Protein
Enzymes)
THE FOLLOWING FORMS THE AF:

1. Amniotic membrane
2. Maternal tissue (interstitial) fluid by diffusion across the amnio-
chorionic membrane
3. Filled from maternal blood
4. Fluid secreted by the fetal respiratory tract
5. Fetal urine
IMPORTANCE OF AF ANALYSIS
Diagnosis genetic and congenital disorders before birth
Detect fetal distress from HDN or from infection
Assess feta lung maturity
Assess the ability of the fetus to survive early delivery
Done during 14 to 18 weeks of gestation or pregnancy

COLLECTION AND PRESERVATION Previous child w/a neural tube disorder (spina bifida or ventral wall
Amniocentesis- needle aspiration of AF from the amniotic sac usually defects)
transabdominal Three or more miscarriages
Maximum of 30 ml AF is collected using sterile syringes
Specimens should be transferred to a sterile tube and taken MACROSCOPIC/PHYSICAL EXAMINATION
immediately to the lab COLOR SIGNIFICANCE
Fluid for bilirubin analysis incases of HDN must be protected from Colorless Normal
the light at all times (most preferred is the Aluminun Foil) Blood streaked Traumatic tap, abdominal trauma intra-amniotic
Fluid for FLM tests should be placed in the ice delivery to the hemorrhage
laboratory and kept refrigerated. Yellow HDN (bilirubin)
Specimens for cytogenetic studies must be kept at room Dark-green Meconium
temperature or body temperature. To prolong the life of the cells. Dark Red-Brown Fetal death
VOLUME
CRYOPRESERVATION
Approximately 35 ml during 1st trimester, peaks during 3rd trimester
After the collection, the AF sample containing he stem cells is
(app. 1L.) & gradually decreased prior to delivery. Major
shipped to the laboratory for processing, cryopreservation, and
contributors are maternal circulation (1st trimester) & fetal urine
storage.
(after the 1st trimester)
Processed sample is exposed to a gradual freezing process which is
POLYHYDRAMNIONS- excessive accumulation of AF usually
important because it keeps the cells alive.
resulting from failure of the fetus to begin swallowing. > 1200 ml.
After freezing, the sample is transferred to a liquid nitrogen storage
caused neural tube disorders, fetal anomalies, chromosomal
tank. Protocols. Used for cryopreservation process have largely
abnormalities.
adapted from those originally designed for bone marrow
OLIGOHYDRAMNIOS- decreased AF due to primarily to increased
hematopoietic stem cells.
fetal swallowing, urinary tract deformities, and membrane leakage,
<800 ml. Caused congenital malformation, umbilical compression,
INDICATIONS FOR PERFORMING AMNIOCENTESIS
death, premature rupture of amniotic membranes.
MATERNAL URINE VS. AMNIOTIC FLUID
F
Maternal Urine Amniotic Fluid
syndrome)
Creatinine 10 mg/dl <3.5 mg/dl
Early pregnancy or child with birth defects
Urea 100 mg /dl 30 mg/dl
Parent is a carrier of a metabolic disorder Glucose & Protein present
History genetic diseases (sickle cell disease, hemophilia sickle cell Fern test for protein Fern like crystals
anemia, muscular dystrophy) &
Sodium Chloride
TEST PERFORMED in AMNIOTIC FLUID Elevated levels indicates anencephaly ( serious birth defect in
1. FETAL FIBRONECTIN (fFN) w/c a baby is born w/out parts of the brain) & Spina bifida (birth
Protein produced during pregnancy. defect involves the incomplete development of the spinal cord
Acts as biologic glue, attaches the fetal sac to the uterine lining or its coverings)
Analyzed when there are symbols of premature labor
Performed if woman is 26-34 weeks pregnant and having 2. ACETYLCHOLINESTERASE/Ache
symptoms of premature labor. Enzyme derived primarily from the neural tissue and is normally
e absent in AF. Its presence in AF in conjuction with elevated AFP
Alerts doctors in stability of preterm delivery values is highly diagnostic of
FETAL DISTRESS TESTING

Most common cause of death in premature newborn. FETAL LUNG MATURITY TESTING
Insufficient lung surfactant packed in lamellar bodies, prevents 1. LECITHIN:SPINGOMYELIN RATIO and PHOSPHATIDYLGLYCEROL
alveoli from collapsing. Major lung surfactant LECITHIN
Surfactant level is corrected with fetal lung maturity & lung Up to 33rd week of gestation the L:S is equal
stability. After 34th weeks gestation lecithin is increased, and
sphingomyelin is decreased
1. HEMOLYTIC DISEASE OF THE NEWBORN/HDN (erythroblastosis Uses anti-phosphatidylglycerol to detect the lung surfactant
fetalis) (lungs require surfactant a soap-like substances to lower the
Maternal antibodies destroy fetal RBC surface pressure of the alveoli in the lungs)
Measurement of AF bilirubin L:S of 2.0 or greater fetal pulmonary system maturity/ normal
Liley graph- semilogarithmic plot of the AF against fetal L:S ratio of <1.5 indicates immaturity relative safe and preterm
gestational age to assign severity of HDN or access fetal risk in delivery
cases of HDN guide the physicians whether to induce laborer or
perform the intrauterine blood transfusion Surfactant
Phospholipid produced by alveolar type II cells
TESTING FOR THE NEURAL TUBE DEFECTS Lowers surface tension
1. FOLIC ACID DEFICIENCY during pregnancy
Glycoprotein produced in early life by the liver & yolk sac of a tension increases
developing baby during pregnancy The half-life of surfactant is 30 hours
Widely used biochemical blood test used to screen for liver,
testicular and ovarian tumor markers.

2. FOAM STABILITY INDEX/ FOAM TEST/ SHAKE TEST CULTURE

Screening test for lung surfactant Evidence of infection is by mounting of the importance of
Used to measure lung surface lipid concentrations microorganism in the AF contributing to the incidence of
AF mixed w/ increasing amounts of 95% ethyl alcohol is shaken pattern delivery spontaneous abortion.
vigorously for 15 secs. & allowed to sit undisturbed for 15 min. Even bacterial vaginosis & trichomoniasis have been linked to
Uninterrupted ring of foam in the tube indicates foam stability preterm birth.
3. AMNIOSTAT-FLM
An immunologic semiquantitative slide agglutination test for
determining the presence of phosphatidylglycerol/PG to detect
the fetal surfactant
Not affected by blood or meconium
4.
Lamellar bodies (lamellated phospholipids that represents a
storage from a surfactants secreted by the type II pneumocytes
of the fetal lung)
Test not affected by hemolyzed blood or meconium
5. OPTICAL DENSITY AT 650 NM

Increased in OD of AF caused by the presence of lamellar bodies
then centrifuging the AF at 2000g for 10 min. & reading the
absorbance at 650 nm
6. MICROVISCOSITY FLUORESCENCE POLARIZATION ASSAY

Provides fluorescence polarization surfactant: albumin ratio
Phospholipids decreases the micro viscosity of AF & the change
is detected in determining the surfactant to albumin ratio

2. CEREBRO SPINAL FLUID (CSF) PURPOSE OF CSF EXAMINATION

Product of continuous secretory activity of the choroid plexuses of Detect bacterial presence
the ventricles of the brain Detect malignant cells
Important in the diagnosis of disorders of the Central Nervous Detect any abnormality affecting in the CNS
System (CNS) Serves as a guide in the diagnosis & prognosis of the diseases of the
LOCATION OF THE FLUID: CNS
1. INTERNALLY- fills the ventricles of the brain, cisternae & the canal of the
spinal cord COLLECTION OF CSF
2. EXTERNALLY- fills the space between the pia & arachnoid membrane TYPES OF PUNCTURE
Lumbar puncture
FORMATION & PHYSIOLOGY Cisternal puncture (cisterna magna or suboccipital)
THE BRAIN & SPINAL CORD- are lined by the meninges, which Ventricular puncture
consist of three layers. Lateral cervical puncture
DURA MATER (hard mother)- outer layer that lines the skull and
vertebral canal. AMOUNT TO BE COLLECTED
ARACHNOID MATER (spiderweb-like)- inner membrane. 8-10 ml, with three sterile labelled, capped tubes numbered 1,2 & 3
PIA MATER (gentle mother)- layer lining of the brain & spinal in order, which they are filled 2 to 7 ml each.
cord. TUBE 1- Chemical & serological test (least affected by blood or
THE ENDOTHELIAL CELLS through out of the body are loosely bacteria)
connected w/c allows passage of soluble nutrients and waste, while TUBE 2- Microbiology Test
CELLS IN THE CHOROID PLEXUSES have very tight-fitting junctures TUBE 3- Cell count & Hematology test
prevent the passages of molecules.
Tight fitting structure in the choroid plexuse -
***NOTE***
FUNCTION OF THE FLUID: TUBE 1 may contain blood from puncture.
For the protection of the brain from injury. Presence of blood affects all the tests except CHLORIDE.
Acts as a medium of exchange for the transfer of dialyzable If the fluid obtained is XANTHOCHROMIC, a trace of lithium oxalate
material. is added to TUBES 2 & 3 to prevent clotting.
Equalizes the pressure between the brain & spinal cord. Cell count, bacterial examination & sugar must be done ASAP,
Serves as excretory channel in the elimination of products of remaining tests can be delayed several hours if the specimens are
nervous metabolism. kept in the refrigerator.
Maintains acid-base balance of CNS. Venous blood should be drawn at the same time as the spinal fluid,
Supplies nutrient to the brain. if the chemical tests are to be done, especially chloride & sugar.

DIFFERENCE BETWEEN TRAUMA & CEREBRAL HEMORRHAGE TAP 4. REACTION- alkaline pH 7.30- 7.45
TRAUMATIC HEMORRHAGE
More blood in the TUBE 1 than Even distribution of blood in 3 5. SPECIFIC GRAVITY- 1.006 TO 1.008
TUBES 2 & 3 tubes
Forms clot Does not form clots 6. PRESSURE
NO XANTHOCHROMIA XANTHOCHROMIC, supernatant, Normal pressure for horizontal position varies between 70 to 200
cytophagocytois, hemosiderin mm. of water (0.8 mm of mercury) with the average of 100 to 150
laden macrophages mm.
CSF pressure is directly related to the pressure in the jugular &
MACROSCOPIC/ PHYSICAL EXAM
vertebral veins, w/c communicate with the intracranial, dural
1. AMOUNT
sinuses & spinal dura.
1 cc/ pound of the body weight
Increased in congestive heart failure, obstruction of the superior
100 to 150 ml everyday production vena cava, straining, breath holding pressure against the abdomen.
10 to 60 ml in children
INCREASED
Acute & chronic congestion of the meninges,
Acute & chronic infections
DECREASED
Circulatory collapse
Severe dehydration
Leakage of CSF
Complete spinal subarachnoid block
2. COLOR CHEMICAL EXAMINATION

Bright Red- due to fresh blood from puncture. 1. CSF PROTEIN
Greenish/ Grayish due to pus cells in severe inflammatory Most frequently performed, varies with age & with the site from
reactions. w/c the specimen was obtained
Yellowish discoloration of CSF (xanthochromia) pale pink to yellow CSF protein 15 to 45 mg/dl
due to RBC lysis & hemoglobin breakdown. Albumin makes up the most of the CSF protein
Prealbumin 2nd most prevalent
3. TRANSPARENCY Alpha globulins (haptoglobins , ceruloplasmin)
Haziness is produced by 200-500 white blood cells / cu.mm Beta globulins (transferrin)
over 500 white blood cells / cu.mm causes TURBIDITY Gamma globulins (IgG, small amount of IgA)

TAU, a separate carbohydrate def transferrin fraction seen in CSF presence in CSF is indicative of recent destruction of the
but not in serum myelin sheath that protects the axons of the neurons
Not found in CSF are IgM , fibrinogen, Beta-lipoprotein. (demyelination)
2. ALBUMIN & IgG 2. CSF GLUCOSE

The CSF/ serum albumin ratio may be used to evaluate blood-CSF Glucose enters the CSF by SELECTIVE TRANSPORT across the BBB,
barrier. which results in a reference value that is approximately
CSF / serum albumin ratio = Albumin CSF g/ml 60% to 70% that of plasma glucose.
Albumin SERUM g/ml HIGH CSF GLUCOSE- results from plasma elevation like diabetes and
Normal ratio is about 1/2305. This ratio is sometimes expressed as in some cases of encephalitis
the CSF/ serum albumin index w/ Albumin CSF in mg/dl & the LOW CSF GLUCOSE- diagnostic value in determining the causative
Albumin SERUM in gms/dl agent in meningitis.
Normal range for this index is about 4 to 8 *LOW CSF GLUCOSE INDICATES BACTERIAL MENINGITIS
Values 9 to 14 interpreted as slight impairment *HIGH WBC COUNT
Values 15 to 30 interpreted as moderate impairment *LARGE PERCENTAGE OF
NEUTROHILS
Values over 30 indicates severe impairment of the blood CSF
barrier.
*LOW CSF GLUCOSE TUBERCULAR MENINGITIS is
Elevated Protein values seen in pathologic conditions *HIGH WBC COUNT suspected
BBB damage (meningitis & hemorrhage)
Immunoglobulins production within the CNS of NEUTROHILS
Decreased normal protein clearance *NORMAL CSF GLUCOSE VIRAL MENINGITIS is suspected
Neural tissue degeneration *INCREASED LYMPHOCYTES
Abnormally low values are present when fluid is leaking from *DECREASED CSF GLUCOSE Primarily caused by alterations in
the CNS the mechanism of the glucose
transport and by increased
METHODS USE IN CSF PROTEIN consumption of glucose by the
Turbidity production by Nephelometry brain cells
DYE BINDING TECHNIQUE
3. CSF LACTATE
PROTEIN FRACTIONS
Normal values= 11 -22 mg/dl
ELECTROPHORESIS
CSF Lactate levels can be valuable aid in diagnosing & managing
To detect oligoclonal bands in the gamma region
meningitis cases.
causing inflammation within the CNS
Primary source is anaerobic metabolism in CNS
MYELIN BASIC PROTEIN
Condition associated with tissue hypoxia of the CNS may cause WHITE CELL <3 >500 <100 100-500
INCREASED CSF Lactate COUNT
Increased in cerebral infarct, traumatic brain injury, hypotension OTHERS 90% PMN MONOCYTES MONOCYTE
> 35 mg/dl seen in BACTERIAL MENINGITIS *10% >90% S
PMN
< 25 mg/dl seen in VIRAL MENINGITIS
*30%>50%
PMN
4. CSF GLUTAMINE
Produced from ammonia & alpha-ketoglurate by the brain cells.
ENZYMES
Serves removes toxic ammonia from the CNS LDH (Lactic DeHydrodenase)- normally present in CSF
Normal value= 8 18 mg/dl
Cholinesterase- no clinical correlation, normally seen in CSF
Elevated levels are associated with liver disorders
PHI (Phosphohexose Isomerase)- increased in brain tumor
GOT (Glutamic Oxaloacetic Transaminase)- increased in cerebral
***NOTE***
infarct & following convulsion
As CSF Ammonia INCREASES the supply of the alpha-ketoglutarate
CK (Creatine Kinase)- normally present in CSF
becomes depleted, the glutamine can no longer be produced to
remove the toxic waste thus patient is suffering in a COMA
OTHER CHEMICAL SUBSTANCES
condition.
Are perform when they are requested by the physician
Requested for coma of unknown origin.
NORMAL BACTERIAL VIRAL FUNGAL or

MICROSCOPIC EXAMINATION
TB
CELL COUNT
PRESSURE 5-20 >30 Normal
(cm H20) /midly 1. If the fluid appears CLEAR LOW CELL COUNT
increased 2. Method for Moderate Cell Count
APPEARANCE Normal Turbid Clear Fibrin web 3. Method for High Cell Count- Differential count
PROTEIN 0.18-0.45 >1 <1 0.1-0.5 4. Method for Counting Mixture of White and Red Cells.
(g/L)
GLUCOSE 2.5-3.5 <2.2 normal 1.6-2.5 CALCULATION:
(mmol/L) WBC (blood) X RBC (CSF)
normal 60-90% normal RBC (blood)
positive = WBC (CSF) corrected
GLUCOSE CSF 0.6 <0.4 >0.6 <0.4
SERUM RATIO 1. CSF CELL COUNT

METHODS OF CELL COUNT

Properly mix the CSF sample
considered normal in NEWBORNS Nine drops of CSF is diluted with one drop of CSF diluting fluid (in
***NOTE*** the ratio 9.1)
Cell counts should be done within 30 minutes after withdrawal of Add 10 ml of glacial acetic acid and 0.2 grams of crystal violet to a
the specimen to avoid disintegration. 100 ml volumetric flask. Dilute to the mark with distilled water
The counting chamber is covered with a cover slip
Charge the counting chamber with fluid and allowed to stand for 5
EXAMPLE: min. for the cells to settle.
Calculation factors when using 1 in 10 C.S.F dil: Cells are counted in all the nine squares
USING AN IMPROVED NEUBAUER CHAMBER GROSS APPEARANCE

* If 64 cells are counted in 4 squares TEST CLEAR HAZY BLOOD CLOUDY BLOODY
No. of Cells X Dilution Factor / Volume TINGED
64 X 10 / 0.4 = 1600/ mm3 0-200 >200 Unknow High Unknown
*Report as 1600 X 106 cells/l mm3 mm3 n
Dilution for None 1:2 turk 1:2 turk 1:30 1:2 turk
USING FUCHS-ROSENTHAL CHAMBER
counting cells soln soln. turk soln soln.
*If 64 cells are counted in 5 squares
64 X 10 / (5 X 0.2)= 640 / mm3 No squares to 9 9 9 9 or 4 9 or 4
count on
hemocytomet
*Report as 640 X 106 cells/l.
er
0-400 Unknow >400 Unknow >6000
STANDARD NEUBAUER CALCULATION FORMULA (CELLS/L) mm3 n mm3 n mm3
Dilution for None None None None 1:200
Number of Cells Counted X Dilutions
counting cells biologic
Number of Cells Counted X Volume of 1 square
saline
= cells/ul
No squares to 9 large 9 large 9 or 4 4 or 5 5 small
Can be used for diluted and undiluted samples count on large large
hemocytomet
er

Cytospin undilute Dilute Straight Dilute Dilute by For Staining- ROMANOWSKY STAIN
dilution d with or by with nucleated -Wright
(0.25 ml/ 5 saline to nucleate saline to cell count; -Wright-Giemsa
drops) 100- 200 d cell 100- 200 if RBC is -
Of fluid mm3 count mm3 counted >1 Also Rapid Staining Methods- DIFF-QUICK
nucleate nucleate million/
d cell d cell mm 3make
DIFFERENTIALS
count count a push
smear and Performed on a stained smear made from CSF
differentiat It is recommended that stained smears be made even when the
e cells that total cell count is within normal limits.
are pushed
out on the Count 100 cells in
end consecutive oil-power
fields
TABLE SHOWING CSF APPEARANCE IN WBC & RBC CELL COUNTS Report percentage
- of each type of cell
present.
Cytospin- this method is designed to concentrate cells that is found
in small numbers
DIFFERENTIAL COUNT TABLE SHOWING DIFFERENT CONDITIONS IN CSF

PLEOCYTOSIS- originally an increased in lymphocytes, now it TABLE 1.
includes the increase of all or any type of cells (especially CONDITION MAJOR GLUCOSE PROTEIN PRESSURE
neutrophils) CELL
(per/ul) TYPE
DIFFERENTIAL COUNT Acute 100- PMN D >100mg/dl I
Indicated when total count is elevated bacterial 100,000
Various methods: meningitis
-Centrifugation- if the total cell count is less than 500 cells/ul Cerebral >500,000 RBC N I I
hemorrhage
-Membrane Filtration- even small number of cell can be examined
(bloody)
-Sedimentation Technique

Gullain- 0-100 L N >100 N FLUOROCHROME SMEAR to detect M. tuberculosis in CSF.

Barre mg/dl
syndrome When facilities for fluorescence microscopy are available,
Multiple 0-50 L N N or I N examination of an auramine stained smear is a more sensitive.
Sclerosis
Spinal Cord 0-50 L N N or I N 2. INDIAN INK
Tumor
One drop of black ink is placed into one drop of CSF and it is
Viral 100- L N N or I N or I
examined using a 40X objective lens on Light Microscope
Infections 2,000
It i
yeast.
*D- decreased The CLEARING is the POLYSACCHARIDE CAPSULE of Cryptoccocus
*I- increased neoformans or C. gattii.
*L- lymphocyte Specificity is improved when you look for budding yeast cell.
*N- normal
*PMN- polymorphonuclear leukocyte
MISCELLANEOUS EXAMINATION
ACID FAST STAIN/ AFB
INDIAN INK PREPARATION
CULTURES
PARASITES
SEROLOGIC EXAM
LIMULUS LYSATE
3. CULTURES
Culture media used are BLOOD AGAR, SABOURAUD MEDIA
1. ACID FAST STAIN / AFB
Colonies are yeast-like, slimy cream to brown in color
Acid staining should be done if tuberculosis is clinically suspected or
the CSF contains lymphocytes and glucose concentration is low and
4. PARASITES
the protein raised.
Amoeba Naegleria fowleri Natural CNS parasite
Only 37% of initial smears will be positive for acid-fast bacilli. This
Tapeworm Taenia solium Natural CNS parasite
result can be increased to 87% if four smears are done.
Protozoa Plasmodium falciparum Natural CNS parasite
Sensitivity also can be increased by examining the CSF sediment.
Protozoa Trypanosoma brucei Natural CNS parasite
Protozoa Toxoplasma gondii Opportunistic CNS parasite

5. SEROLOGIC EXAMINATION 7. GRAM STAIN

Serologic examination of CSF is typically involved with diagnosis of
NEUROSYPHILIS the doctor suspects the patient have meningitis or encephalitis
A fluorescent treponema anticody absorption (FTA-ABS) test for CSF caused by bacteria or fungi.
has been developed. Meningitis is an inflammation of the membranes surrounding the
The VENEREAL DISEASE RESEARCH LABORATORY (VDRL) test is also spinal cord and brain. It progresses quickly and can be life-
used. threatening in some cases.
LATEX AGGLUTINATION (LA) allows rapid detection of bacterial Organisms includes Meningoccoci, Pneumococi & Hemophilus
antigens in CSF influenza in young children
Tests are available to detect N. meningitis group A, B, C, Y and W135
(Group B reagent cross-reacts with E. coli K1 antigen)
H. influenza type B, S. pneumonia, and S. agalactiae.
Sensitivity varies greatly between bacteria L.A for Haemophilus
influenza has a sensitivity of 60 to 100 % but is much lower for other
bacteria.
6. LIMULUS AMEBOCYTE LYSATE

LIMULUS amebocyte LYSATE- test is a simple and cost effective
means to screen CSF (cerebrospinal fluid) for Gram- negative agents
of meningitis.
DIAGNOSIS:
LIMULUS AMEBOCYTE LYSATE (L.A.L) an aqueous extract of blood
cells (amoebocytes) from the horse shoe crab, (Litmus polyphemus)
LAL reacts with bacterial endotoxin or lipopolysaccharide (LPS),
POLYMERASE CHAIN REACTION (PCR) is a technique used to

amplify small traces of bacterial DNA.

3. GASTRIC FLUID SEVERAL SUBSTANCES SECRETED BY THE STOMACH NEEDED FOR PROPER
DIGESTION
1. HYDROCHLORIC ACID - secreted by parietal cells hydrolyzes peptides

IMPORTANCE OF GASTRIC FLUID ANALYSIS
and disaccharides, and converts pepsinogen to pepsin. Intrinsic factor
Determines whether a patient can secrete any gastric fluid also secreted by parietal cells is needed for the absorption of vitamin
Measures amount of gastric acid that can be secreted by a patient B12
with symptoms of peptic ulcer 2. PEPSINOGEN - secreted by peptic cells catalyzes the degradation of
Determines numerous conditions that disturb functions of the proteins to proteases and peptones
stomach, duodenum, liver, pancreas, and the systemic condition of
anemia 3. Other enzymes secreted by the stomach include peptidase, lactase, and
Helps the physician in the diagnosis of gastric diseases and assist lipase
him in the selection of therapy and prognosis
4. MUCUS - secreted by goblet cells and mucus glands acts to protect the
GASTROINTESTINAL TRACT PHYSIOLOGY
stomach wall from acids and enzymatic activity
Digestive System is composed of the alimentary canal, a continuous 5. CHYME - mixture of digestive secretions and partially digested food (ex.
tube from the mouth to the anus including the esophagus, the Ingested food and water in the stomach and small intestines)
stomach, the intestines, the colon, and the accessory digestive
organs including the mouth, much of the pharynx, the teeth, 6. Gastric secretions are stimulated by neural response sight, smell, or
tongue, salivary glands, liver, gallbladder, and pancreas anticipation of food, distension of stomach by food of fluid, gastric
Contains and processes food from ingestion through digestion and mucosal contact with secretagogues (protein breakdown products)
elimination
7. GASTRIN - hormone secreted by gastric mucosa
DISORDERS OF GASTROINTESTINAL TRACT
A. UPPER GASTROINTESTINAL BLEEDING (UGIB)
bleeding in the esophagus, stomach, or duodenum,

most common gastroenterological emergency
leads to anemia
Diagnostic Test includes:
Repeated hemoglobin levels and occult blood

Fecal Occult Blood - (nonvisible blood in the feces) not as increase blood loss in the GI tract is detected by occult blood test
sensitive as are tests for gastric occult blood (nonvisible blood (hidden or small amounts of fecal blood often not visible to
in gastric juices the naked eye)
B. DIARRHEA COLLECTION OF GASTRIC FLUID
common disorder of the GI tract Collection of the Specimen

frequency and volume of bowel movements are increased and the
The patient must be in a fasting state for 12 hours
BM are more liquid
Contamination with saliva neutralizes the gastric acidity
associated with infectious agents, toxins, malabsoprtion, and a
therefore it should be prevented
variety of GI disturbances
Time specimen should be collected for the purpose of
Caused by large volume of fluid being presented to the large
comparison
intestine:
Due to increased secretions Two Types of Gastric Juice Collection

Due to increased amounts of osmotically active substances
Using an evacuated tubes:
remaining in the GI tract
Because of hypermotility (increased intestinal movement) 1. Levine Tube - inserted in the nose (nasal intubation)
resulting in decreased intestinal absorption 2. Rehfuss Tube - inserted in the mouth (oral intubation)
results in dehydration and critical electrolyte imbalances
Two Laboratory Methods for Gastric Collection
C. MALABSORPTION
1. Intubation Technique - allows measurement of GF using
state of abnormal digestion or absorption of a single nutrient or of quantitative evaluation
multiple nutrients through the GI tract that may lead to
malnutrition or anemia
fats, meats, fibers, and carbohydrates all may be improperly
digested with malabsorption
Maldigestion is impaired digestion caused by lack of digestive
enzymes (trypsin and chymotrypsin)
D. COLORECTAL CANCER
common cancer of the GI tract

A. NASOGASTRIC INTUBATION H+ in gastric secretion combines with the resin and liberate
tube is carefully inserted through the nasal passage, down the azure-A ions, which are then absorbed into the
esophagus, and into the stomach bloodstream in the small intestine and subsequently
excreted in the urine
LABORATORY EXAM OF GF
GROSS/PHYSICAL EXAMINATION
1. VOLUME
20 - 100 mL
2. Tubeless Technique - investigates the ability of the stomach to - gastric hypomotility, Zollinger-Ellison Syndrome
produce acid - gastric hypermotility, Pernicious Anemia
TUBELESS GASTRIC ANALYSIS 2. COLOR
(A) Normal appearance is a translucent pale gray color
gastric secretion is stimulated and after one hour, a dye bound may contain spots of blood
to resin (Azure A) is given orally brown, green, or yellow - due the reflux of bile during the intubation
based on the surrounding pH, the resin releases the dye process
It is absorbed and excreted in the urine 3. ODOR
the quantity of dye in urine indicates the acidity of gastric juice.
It rules out achlorhydria Odorless or slightly sour or faintly pungent
used as a screening test Abnormal Odors:
(B) Fecal Odor - seen in intestinal obstruction or gastrofolic-
use of an orally administered ion-exchange resin to detect the fistula
presence of H+ in gastric fluid Foul-Putrid - seen in carcinomatous ulcer
marketed under the trade name, DIAGNEX BLUE
based on the use of a carboxyl-acid cationic resin (amberlite XE- Alcoholic Odor- seen in alcoholic coma or after alchol test
96) with an innocuous indicator dye, Azure A coupled to meal
it Ammoniacal Odor - seen in cases of anemia

Rancid Odor - due to butyric (fatty acid) and actic acid C. FREE ACIDITY - Free HCl in the free state
(present in milk) indicating stenosis and fermentation
(+ violet color with bluish tinge)
4. REACTION
D. COMDINED ACIDITY
Acidic, pH 1.6-1.9
Total Acidity - Free Acidity = Combined Acidity
High acidity - pH of 1.4 or lower
2. LACTIC ACID
Low acidity - pH of 2..0 to 2.8
Achlorhydria/Anacidity - pH 3.0 or higher Not normally present in GF, positive after ingesting sour milk
Neutral or alkine - due to contamination with saliva formed from fermentation of carbohydrates by lactic acid forming
bacteria, the Boas-Oppler bacillus (Lactobacillus acidophilus)
5. SPECIFIC GRAVITY suggests advanced carcinoma of the stomach
1.001 to 1.010 (ave. 1.007) 3. OCCULT BLOOD
CHEMICAL EXAMINATION - specialized screening tests are used for the detection of occult
1. ACIDITY blood in gastric aspirate or vomitus (Gastroccult Slide Test) ; +blue color
A. TOTAL ACIDITY - includes free HCl and the combined acids 4. ENZYME TEST
TESTS FOR TOTAL ACIDITY: a) Rennin - responsible for the curdling of milk, if absent
indicates organic disease of the stomach
a) Phenolphthalein Test (+ deep pink color)
b) Pepsin - secreted by the chief cells of the stomach, acts as
b) - for Total Acidity and free HCl protein when activated by HCl
(+ red end point, salmon pink) MICROSCOPIC EXAMINATION
B. FREE HCl - HCl not combined with proteins Gastric Fluid usually separates into 3 layers:
TESTS FOR FREE HCl: Top layer - mucus
a) Top Central Layer - Opalescent Fluid
b) (+ rose-red color) Bottom Layer - Sediments
c) (+ purplish-red color) Make a smear of the unfiltered gas
staining. Can be found are cells found in the lining of the stomach and
digestive tract:
1. EPITHELIAL CELLS - squamous cells from the esophagus has no 4. FECAL ANALYSIS
significance. Columnar cells indicates gallbladder disease.
Provides important information related to gastrointestinal
2. WHITE BLOOD CELLS - seen in abcess of the stomach and disorders, infections, and several other medical conditions
gastric carcinoma, inflammatory condition.
After chyme has remained in the large intestines for 3-10 hours, it
3. RED BLOOD CELLS - seem in irritation of the gastric mucosa normally becomes solid or semisolid and is then called feces
4. MUCUS - occasionally seen as small spherical snail-like bodies GROSS EXAMINATION
suggests hypertropic gastritis ulcer
Appearance of feces provides some clues to possible GI disorder
5. BACTERIA - seen in the absence of Free HCl, usually seen are
the gram positive bacteria like Boas-Oppler bacilli, TB bacilli, Hematochezia - bright red blood in feces
and sarcinae Acholic Stool - very pale stool indicates biliary obstruction
6. FOOD PARTICLES AND RESIDUE - comes from test meals and Melena - black, tarry colored stool seen in upper GI tract bleeding
stimulants that are given.
7. YEAST - suggest retention and fermentation, few may be MICROSCOPIC EXAMINATION

seen.
Fecal Leukocytes - esp. Neutrophils, are commonly associated with
8. TISSUE FRAGMENTS - seen in cancer, necrotic tissue dysentery or invasion of the intestinal wall
containing WBC and bacteria
Eosinophils are also present in amebic infections
TERMS RELATED TO ACIDIMETRY
ACHLYIA - complete absence of HCl CHEMICAL EXAMINATION

EUCHLORHYDRIA - normal acidity pH 1.6-1.8 A. Fecal Occult Blood
HYPOCHLORHYDRIA - decreased acidity Fecal blood is found in infection, in trauma, and in colorectal cancer
Early diagnosis is associated with a goof prognosis which is
HYPERCHLORHYDRIA - increased acidity
beneficial in the early diagnosis
ACHLORHYDRIA - absence of acidity Positive occult blood test is caused by ulcers, hemorrhoids,
inflammatory conditions and infectious agents

THREE TYPES OF FOBTs C. Fecal Fat Testing
each with a different part of hemoglobin molecule conditions that decreases the production of pancreatic enzymes
such as cystic fibrosis, pancreatic insufficiency, pancreatitis, and
Guaiac Test - most common method for detection of fecal blood
pancreatic carcinoma as well as the absence of the bile salt that
assist lipases in fat breakdown, cause steatorrhea (an increase in
Example:
Is hemoccult require the peroxidase-like activity of fecal fat)
intact heme. If hemoglobin or another peroxidase or floats in water, pale and greasy oily appearance and a foul smell
psuedoperoxidase is present in the feces in sufficient before any fecal fat testing, it is important for patient to have a diet
amounts, the quaiac paper will develop turns blue with a normal amount of fat intake
when hydrogen peroxide developer is applied Mineral oils and many creams can cause false positive result
Immunologic- Hemoccult-ICT react with antigenic sequences on
based Test human globin chains. More specific for lower GI
bleeding. The test can not be used on GF specimens D. Qualitative Fecal Fat Testing
because globin proteins may be completely destroyed
by peptidase of the proximal gut are performed microscopically for triglycerides (neutral fats), fatty
Heme-Porphyrin- Less affected by the anatomic site of bleeding. acids salts, fatty acids and cholesterol stained using Sudan III. Sudan
Bases Test Porphyrin ring remains intact during transit and fecal IV, and Oil Red O
storage. Fluorometric assays based on the conversion number and size of fat globules (neutral fat) are detected
of non-fluorescing heme to fluorescent porphyrins qualitatively by staining with Sudan III in 95% ethanol in wet
preparation
B. Fetal Hemoglobin (APT TEST) E. Quantitative Fecal Fat Testing
Newborns may excrete stools or vomitus containing blood, which to follow up a positive qualitative test
originate from maternal blood ingested ate delivery or from the Dietary requirements - requires adherence to a diet of 100g/day
er the hemoglobin present is fat intake before and during test collection
hemoglobin A (maternal or hemoglobin F (fetal) in origin chemistry department usually sent to reference laboratory
Stool or vomitus is mixed with water to yield a pink supernatant is 3-day (72-hour) collection Test Method
removed then alkalinized with dilute NaOH. If the pink color Van de Kamer - classical titration. Uses sodium hyroxide to
remains, the blood contains fetal hgb. If the pink color changes to chemically titrate the amount of fat
yellow or brown, the hgb in the sample is maternal hgb. Acid steatocrit
Near infra-red spectroscopy

FECAL ENZYMES 5. SEROUS FLUID
Pancreatic insufficiency and cystic fibrosis Serum-like composition

Lungs, heart and abdominal organs are surrounded by a thin,
Pancreatic-associated enzymes
continuous membrane, as well as by the internal surfaces of the
Tryspin-Classical Tryosin Test - series of diluted stool specimens are body cavity wall:
placed on x-ray paper (has a gelatin coating). After incubation, the stool Visceral membrane space or cavity filled with fluid lies
is rinsed off and the paper evaluated to determine the dilution at which between the membrane that covers the organ
no gelatin has been digested by the protease trypsin. Test not sensitive Parietal membrane lines body wall
Chymotrypsin - more sensitive and can be measure Lungs are individually surrounded by a pleural cavity, heart by
spectrophotometrically pericardial cavity and abdominal organs by peritoneal cavity
Effusion accumulation of serous fluid
Elastase I - pancreas speicific enzymes not affected by motility or ther
mucosal issues. Immunoassay procedure provides higher degree of Ultrafiltrate of plasma and maintained by pressure forces (tissue
specificity colloidal osmotic pressure, capillary hydrostatic pressure, capillary
colloidal osmotic pressure and tissue hydrostatic pressure) and by
absorption of fluid into lymphatic system
Result from a disruption in the balance of pressures (Transudates)
or in response to infection and inflammatory processes (Exudates).
Correct classifying effusions assists physicians to determine a
diagnosis. Classifications are based on results from various
laboratory tests
LABORATORY TESTING OF SEROUS FLUIDS
MACROSCOPIC EXAM
Clear and pale yellow - Normal
Sanguineous haemorrhage but also appear similar in traumatic
taps
Purulent presence of WBCs which correlates with inflammation or
infection
Milky contain chyle or pseudochylous
Shimmery golden-green iridescence with cholesterol crystals seen
Brownish contain mixture of RBC and WBC along with lining cells measurement combined with fluid cytologic exam, identification
or malignant cells of malignant effusions is significantly increased
Clotted reported as fibrinous
MICROSCOPIC EXAM
CHEMICAL EXAM RBCs not normally seen, if present may indicate haemorrhage or
Standard chemical tests perform include glucose, LDH and Protein traumatic collection
Used to categorize effusions as transudates or exudates WBCs normally present in low numbers with mononuclear cells
Less commonly performed: Alkaline phosphatase, ammonia, predominating. Presence of increased number of WBCs correlates
amylase, bilirubin, chloride, lipids and pH with pathologies
1. Peritoneal fluid ALP increased Mesothelial cells normal sloughing wof cells, may exhibit reactive
small intestine perforated morphology that can be confused with plasmacytes, histiocytes or
2. Peritoneal fluid NH4 levels tumor cells
higher than serum levels in cases of bowel strangulation,
perforated peptic ulcer, ruptured appendix and bladder MICROBIOLOGICAL EXAM
3. NH4 and Amylase Gram stain on both aerobic and anaerobic cultures should be set uo
Increased in bowel necrosis to increase rate of microbial recovery
4. Amylase Acid Fast Stain on pleural fluids should be routinely performed for
Increased esophageal perforation, metastatic adenocarcinoma, TB
pancreatitis Fungal stains and cultures done if yeast infection is suspected
5. Chloride
Levels of body fluids < serum when bacterial infection is present TRANSUDATES VS EXUDATES
with WBCs Transudates Effusion
6. Lipids testing Occur during systemic disorders that disrupt fluid filtration and fluid
Assists in differentiation between chylous (TAGs) and absorption which includes CHF, hepatic cirrhosis, nephrotic
pseudochylous (Chol) effusions syndrome
7. Low pH value
In pleural fluid, can help identify patients with parapneumona Exudate Effusion
effusions that require aggressive treatment Occur during inflammatory processes that result in damage to blood
8. Carcinoembryonic Antigen (CEA) vessel walls, body cavity membrane damage, decreased
Tumor marker useful in evaluating pleural and peritoneal reabsorption by lymphatic system
effusions from patients who have previous history or currently Includes infection, inflammation, hemorrhages and malignancies
suspected of having CEA-producing tumor. When CEA Can damage tissues, body cavity membranes and alter lymphatics
DIFFERENCE BETWEEN TRANSUDATES AND EXUDATES TYPES OF SEROUS FLUIDS

PARAMETERS TRANSUDATES EXUDATES Found in cavities surrounding vital organs
ORIGIN Non-inflammatory Inflammatory 1. PERICARDIAL FLUID
PH Alkaline Acidic Accumulation of fluid around heart
SG <1.018 >1.018 Normally contains <50ml of fluid
COAGULATION No clot formation Clot formation (due to
Pericardiocentesis removing excess pericardial fluid. Dangerous
fibrinogen)
PROTEIN <3gm >3gm and rarely performed
GLUCOSE Same with blood Lower than blood Normal fluid pale yellow and clear
glucose glucose Sanguineous (blood) present due to number of causes
LDH 200 IU/L >200 IU/L Caused by damage to mesothelium
CHLORIDE Lower than blood Higher than blood Always EXCUDATES
chloride glucose
CELLS Less (usually Many
2. PLEURAL FLUID
lymphocyte) (neutrophils/PMNs)
Accumulation of fluid around the lungs
CRYSTALS Absent Present
Normal amount is <30ml. abnormal accumulation begins at the base
CHYLOUS EFFUSION of the lungs
Effusion that contains emulsion of lymph and chylomicrons Thoracentesis performed to remove excess fluid from pleural sac
Obstruction or damage of lymphatic vessel contributes to the Removal of fluid not only provides specimen for lab examination but
development of a chylous effusion also helps improve patient symptoms and allow for better
Appear milky and may appear shimmery (resembling milk mixed visualization of lungs and pleural cavity during radiologic procedures
with honey) if cholesterol crystals are present in ruptured lymphatic
vessel 3. PERITONEAL FLUID
Accumulation of peritoneal fluid also called Ascites in the abdominal
CHRONIC EFFUSION cavity
Present in disorders as rheumatoid arthritis and TB May accumulate in abdomen as a result of clinical disorders or
Resemble chylous effusion because of high amount of cellular debris generalized edema
and cholesterol present Paracentesis removal of ascites in abdomen
Termed pseudochylous and can be differentiated from chylous Peritoneal lavage procedure to collect fluid when patient has had
effusions using pH and lipid analysis a blunt or penetrating abdominal trauma
Removal of more than 1000ml ascites can cause hypovolemia and
shock

CYTOLOGIC EXAMINATION 6. SEMINAL FLUID

When malignant disease is suspected, large volume (10 to 200ml) of
Also called: SEMEN or SEMINAL PLASMA
pleural, pericardial or peritoneal effusion should be submitted
Mixture of secretion of several glands in the male reproductive
Cells should be concentrated to increase yield of cells and a cell
organ
block as well as cytocentrifuged smears can be prepared
Fluid that is ejaculated at the time of orgasm
Important, sensitive and specific procedure in diagnosis of primary
Acts as vehicle and a nutrient support medium for spermatozoa
and metastatic neoplasm
Performed by cytologist or pathologist Composed of:
Spermatozoa in their nutrient plasma
Secretions from:
Prostate (20%)
Seminal vesicle (60%)
Testis (5%)
Epididymis (10-15%)
SEMEN PRODUCTION
STRUCTURE FUNCTIONS
Seminiferous tubules of testes Spermatogenesis
Epididymis Sperm maturation
Ductus deferens Propel sperm to ejaculatory ducts
Seminal vesicles Provide nutrients for sperm and
fluid
Prostate gland Provide enzymes and proteins for
coagulation and liquefaction
Bulbourethral glands Add alkaline mucus to neutralize
FRACTIONS OF SEMEN
First Clear, viscid fluid from the urethral and bulbo-
urethral glands
May cleanse and lubricate the urethra in
preparation for the bulk of the ejaculate which
is to follow

Second Consists largely of prostatic secretion along Date and exact time of collection must be noted
with most of the spermatozoa and small For insemination purposes, it can be preserved in frozen state and
amount of secretion from the epididymis stored at 7 degrees Celsius
Final or Third Consists almost entirely of a mucoid secretion
resulting from emptying of the seminal
PHYSICAL EXAMINATION
vesicles
AMOUNT: 3-5ml
Mixing of the various fractions in ejaculation is important to the
proper conduct of the semen examination Volume of ejaculated semen should normally be >2ml. it is
measured after the sample has liquefied. Volume <2.0ml is
SEMENURIA or SPERMATURIA abnormal and associated with low sperm count
Presence or discharge of semen in urine COLOR and TURBIDITY: opaque, pearly white or grayish white
coagulum, thick and sticky
OCCASIONS WHEREIN SEMEN EXAMINATION IS REQUESTED: ODOR: distinct musty or acrid
Examination of male fertility VISCOSITY: fresh ejaculates forms a gel but it liquefies after 10 to 30
Examination of stain in suspected rape minutes (complete liquefaction). Reported as 0 (watery) to 4 (gel-
To check the effectiveness of previous vasectomy like)
Support or disprove a denial of paternity After ejaculation, normal semen is thick and viscous. Becomes
Screen donor for artificial insemination programs liquefied within 30 minutes by the action of proteolytic enzymes
Evaluate semen quality for sperm and semen banking secreted by prostate. If liquefaction does not occur within 60
minutes, abnormal. Viscosity of the sample is assessed by filling a
COLLECTION pipette with semen and allowing it to flow back into the container.
4-7 days period of abstinence from sexual activity is recommended Normal semen will fall drop by
more than 2cm long, viscosity is increased. Increased semen
METHODS: viscosity affects sperm motility and leas to poor invasion of cervical
Masturbation/self-ejaculation or production mucus; it results from infection of seminal vesicles or prostate
During or after coitus interruptus/sexual contact or intercourse REACTION: mildly alkaline ph 7.2-7.8
Aspiration of seminal fluid from the vaginal vault Drop of liquefied semen is spread on pH paper (of pH range 6.4-8.0)
Condom and pH is recorded after 30 seconds. Normal pH is 7.2 to 8.0 after 1
hour of ejaculation.
PRECAUTIONS IN TRANSPORTING SPECIMENS TO THE LABORATORY: The portion of semen contributed by seminal vesicles is basic, while
Specimen must be examined within 2-3 hours after collection portion from the prostate is acidic. Low pH (<7.0) with absence of
Specimen must not be exposed to hot and cold temperature sperms (azoospermia) suggests obstruction of ejaculatory ducts or

absence of vas deferens. Low pH is usually associated with low

semen volume (as most of the volume is supplied by seminal 2.
vesicles). 1 drop semen + 1 drop 0.5% yellow aqueous Eosin stain on glass
SPECIFIC GRAVITY: 1.035 slide
Living will not accept dye, dead ones will stain pink yellow.
3 STAGE PROCESS IN COAGULATION AND LIQUEFACTION Estimation of motility and morphology of spermatozoa are most
1. Formation of fibrin clot by the action of a prostatic clotting enzyme closely related with fertility.
on a fibrinogen-like precursor formed by thr seminal vesicles. Motility is graded as:
2. Liquefaction is initiated by the action of fibrinolytic enzymes of 0 none
prostatic origin 1 poor
3. Fibrin fragments are degraded further to free amino acids and 2 moderate
ammonia y the action of several poorly characterized proteolytic 3 good
enzymes, including an aminopeptidase and pepsin. 4 excellent
MICROSCOPIC EXAMINATION MORPHOLOGY:

MOTILITY: 70-80% motile WHO criteria 5 ul semen + stain on slide, apply coverslip
GRADING Examine at least 200 spermatozoa under OIO. Percentage of
4.0 Rapid, straight line motility abnormal forms should be reported. A specimen that contains <30%
3.0 Slower speed, some lateral movement
abnormal forms is considered normal.
2.0 Slow forward progression, noticeable lateral movement
1.0 No forward progression Head relatively large, oval, flattened, 3-6 microns in length, 2-3
0 No movement microns in width
Movements may be described as quick, sluggish, active and rapid Mid portion or neck small connecting piece between the head
and the tail
MOTILITY TESTS: Tail long, slender part, exhibits violent whip-like motion
This describes the percentage of sperm that are moving. 40% or Entire sperm cell measure 50-70 microns in length, with tail
more of the sperm should be moving composing about 90%
1. REVITALIZATION TEST Abnormalities:
1 drop semen on slide Head too large, too small, absent, double, abnormal
If spermatozoa appears immobile or dead) add 1 drop Glucose shape, atypical distribution of chromatin, swollen and
irregular
Observe whether spermatozoa recover and become motile again

Mid portion or neck absent, swollen, rudimentary, DILUTING FLUIDS

abnormal shape Cold distilled water
Tail rudimentary, absent, double, swollen, too short 1% formalin
5% Sodium bicarbonate
STAINS 0.5% Chlorazane or Chloramine T
Papanicolau 1% Formalin in 3% Trisodium citrate
5% Sodium bicarbonate in 1% phenol
Hematoxylin-Eosin CHEMICAL EXAMINATION

Basic Fuchsin Fructose level main sugar found in human semen. Specimen can
Crystal Violet be screened for the presence of fructose using Resorcinol Test that
Methylene blue produces an orange color. Should be tested within 2 hours of
Viability of spermatozoa is a valuable quality parameter which is collection or frozen to prevent fructolysis. A normal quantitative
measured by permeability of the plasma membrane to specific level of fructose is equal or equivalent than 13umol per ejaculate
stains. The most common staining procedure has been Leukocyte Esterase presence of WBC
eosin/nigrosin being easy to use, requiring only a basic microscope. Acid phosphatase = 2220-2500 King Armstrong units, main enzyme
Its drawback is that assessment of viability can be extremely in seminal and prostatic fluids; is determined on vaginal clothing
subjective due to varying intensity of background coloration. Dead samples, presence indicate recent sexual intercourse. Level of
sperm is red staining and alive sperm is colorless (white). 58% or >50U/sample is considered positive
more of the sperm should be alive. Hormonal evaluation
Hormones measured in evaluation of Male Infertility.
SPERM COUNT Play a critical role in spermatogenesis. Testis has 2 primary
Normal value = 20M/ml to 160M/ml functions:
Neaubauer hemocytometer 1. Testosterone production
Dilute specimen 1:20 using WBC pipet, allow to satnd for 2 2. Sperm production
minutes after charging Both under hormonal control by the brain and there is an
a. Using 5 RBC squares expected relationship between the hormone levels secreted
count x 1,000,000 = sperms.ml by the brain and secreted by the testis
b. Using 2 WBC squares Problems with either function can be detected by blood
tests which can help localize if the brain, testis and/or
or count x 100,000 = sperms/ml

reproductive tract (pipes which carry sperm from testis to head of sperm, they will prevent penetration of egg by sperm. If
penis) is the problem. antibodies are bound to tail, will retard motility.
Basic hormone evaluation includes: Double-fluorochrome soerm-cytotoxic antibody
Testosterone ELISA
Follicle Stimulating Hormone (FSH) Mixed antiglobulin reaction
Prolactin Immunobead test
Optional: Estradiol, Thyroid stimulating hormone
Homonal evaluation is mandatory in all males with infertility What are Antisperm Antibodies?
as a treatable cause of a low sperm density, such as Fight sperm
pituitary adenoma causing low gonado-trophins, may be Happens when immune system mistakenly targets sperm
found; testosterone deficiency may be associated with
impaired spermatogenesis and it can be easily treated even make it harder to have a baby.
if fertility cannot be restored.
Sperm agglutination
IMMUNOLOGIC EXAMINATION Sticking together of sperm
Immunological test done on seminal fluid include mixed antiglobulin Usually measured as a percentage of sperm in a sample that has
reaction (MAR test) and immunological test. Antibodies against become stuck together
sperms immobilize or kill them, preventing passage through cervix Cannot swim properly and will be unable to fertilize an egg
to ovum. Antibodies can be tested in serum, seminal fluid, cervical
mucus 2. IMMUNOBEAD TEST
Antibodies bound to surface of spermatozoa can be detected by
1. SPERMAGGLUTINATING ANTIBODIES (SAA) antibodies attached to immunobeads (plastic particles with
Male antisperm antibodies test attached anti-human immunoglobulin that may be either IgG, IgA or
Gelatin agglutination test IgM
Sperm immobilization test test for antispem antibodies as Percentage of motile spermatozoa with attached two or more
cause of infertility, based on loss of ability of spermatozoa immunobeads are counted amongst 200 motile spermatozoa.
with surface antibodies to move when complement is >50% with attached beads is abnormal
present For wo
Antibodies against sperms immobilize or kill them, preventing pregnant is intrauterine insemination, when doctor puts sperm
passage through cervix to ovum. Antibodies can be tested in serum, directly in uterus, allowing sperm to avoid contact with antisperm
seminal fluid or cervical mucus. If antibodies are present bound to antibodies in cervical mucus

MISCELLANEOUS TESTS 2. Azoospermia

MEDICO-LEGAL CASES identification of semen or proof of presence Total or complete absence of spermatozoa in semen or
1. Florence Test failure of formation of spermatozoa
For choline substance 3. Oligospermia
(+) brown plates or needle like crystal, rhombic periodide of Deficient or decrease number of spermatozoa or only few
choline form motile sperm cells present
2.
Spermine substance
(+) yellow crystals, leaf-like shaped with irregular borders
3. Hyperimmune Sera Test or Precipitin Test
Semen of human origin
(+) formation and deposit of precipitate
SPERM FUNCTION TESTS

1. Hamster egg penetration
Sperms incubated with species nonspecific hamster eggs
and penetration is observed microscopically
2. Cervical mucus penetration
midcyle cervical mucus

3. Hypo-osmotic swelling
Sperm exposed to low sodium concentrations are evaluated
for membrane integrity and sperm viability
4. In vitro acrosome reaction
Evaluation of acrosome to produce enzymes essential for
ovum penetration
CONDITIONS WHICH LEADS TO STERILITY:

1. Necrospermia/Necrozoospermia
Spermatozoa of semen are dead or motionless

EXAMINATION FOR PRESENCE OF SEMEN IN MEDICO-LEGAL CASES 7. SPUTUM

Colposcope examination of material obtained from vagina, stains from
clothing, skin, hair or other body parts from semen.
FORMATION OF SPUTUM
Carried out in cases of alleged rape or sexual assault
Coughed up from the alveoli, trachea, and bronchi of pulmonary
tract
Exudate which is formed in the course of pulmonary, bronchial, or
tracheal infections
Produced by the surface epithelial cells and submucous glands
COMPOSITION OF SPUTUM
Composed of secretions, exudations, and exfoliations of the

bronchopulmonary tree
Normal conditions: mucous secretions of the goblet cells and
mucous glands associated with respiratory epithelium are carried by
ciliated epithelium up to the trachea.
CHEMICAL COMPOSITION
95% Water and 5% Organic Components

Carbohydrates (sialic acid which contributes to its viscosity
Proteins (Immunoglobulins), glycoproteins (Contributes to
its viscoelastic property)
Lipids, enzymes, macrophages, bronchial epithelial cells
COLLECTION OF SPUTUM SAMPLE
Prior to collection, the patient is advised to first brush his teeth,

rinse the mouth very well. Cough up the sputum from down deep
the chest into a clean, dry wide mouth container, cap tightly. Avoid
saliva when during collection.
Early morning specimen is preferred because there is greater output
representing the pulmonary secretions accumulated during the
night.

Specimen for culture should be fresh, placed in a clean cap without Volume
preservative. A 24-hour sputum is measured in Chronic bronchitis, lung
If unable to cough, induction of sputum can be done by heat aerosol abscess, bronchial asthma. An increasing volume of sputum
technique (15% NaCl aerosol spray and propylene glycol for 20 indicates bad prognosis
minutes) Color
PHYSICAL EXAMINATION OF SPUTUM Clear and Colorless Normal

Yellowish Infectious process like Pneumonia
pH of sputum Greenish tint Pseudomonas
Normally Alkaline Rust colored Pneumococcal pneumonia
Quantity Bright red Pulmonary infarction, TB,
Morning Sample 2 to 5 mL Malignancy
24-hour collection 100 mL
OTHER COLOS OF SPUTUM
1. 24-hour collection >100mL
Pulmonary edema
Bronchiectasis
Lung abscess
Advanced Pulmonary TB
2. >500 mL
Rupture or amoebic liver abscess into lung
MACROSCOPIC EXAMINATION
Reaction
pH 6.5 7.0
May be alkaline in cases of Chronic inflammation and
necrosis
Specific Gravity
Consistency
MUCOID 1.004 1.008
Bloody
PURULENT 1.015 1.060
Rusty Colored
SEROUS >1.037
Purulent
Foamy white
Frothy pink

CONSISTENCY AND APPEARANCE MICROSCOPIC EXAM OF SPUTUM
Serous Pick out cheesy masses, purulent particles, rusty portions and
Frothy suspicious structures
Colorless or yellow fluid in pulmonary edema
2 Kinds of Smears
Mucous
Glassy 1. Stained Preparation
Tenacious found in: AFB
acute Bronchitis
Asthma Giemsa
Lobar Pneumonia 2. Unstained Preparation
Whooping cough Examine for;
Purulent
Ruptured empyema crystals
Bronchiectasis parasite using the direct smear
Tenacious
MICROSCOPIC FINDINGS
Thick and viscous due to mucus found in:
Lobar Pneumonia Caseous masses
Bronchiectasis
SPUTUM VS. SALIVA s Asthma

Charcot-Leyden Crystals Asthma
Criteria Sputum Saliva Bronchial casts
Color White/Colorless Colorless Concretions
Density Densest Broncholith
Appearance Viscose ll out Calcified particles as seen in Broncholithiasis
MICROSCOPY
Lung cancer cells
Epithelial Cells < 25/LPF > 25/LPF
Central bronchus tumors
WBCs > 5/LPF < 5/LPF
May require 4 samples to detect
Food Particles (None/Few)/LPF Many/LPF
Bacteria (Few/Moderate)/HPF (Abundant/many)/HPF
Allergy
Asthma

Heart Failure Cells (Hemosiderin Laden Macrophage)
Are siderophages (hemosiderin containing macrophages)

generated in the alveoli of patients with left heart failure or
chronic pulmonary edema, when the high pulmonary blood
pressure causes red cells to pass through the vascular wall.
BLOOD IN SPUTUM (HEMOPTYSIS)
TB
Bronchiectasis
Mitral Stenosis
Pneumonia
Lung abscess
Bronchogenic carcinoma
Charcot-Leyden Crystals
Are formed from the breakdown of eosinophils and may be seen in

the stool or sputum of patients with parasitic diseases.
These crystals can appear in a variety of sizes and only indicate an
immune response, but the cause may or may not be a parasitic
infection
Crystalloids containing galectin-10 (Eosin lysophospholipase binding
protein)
A microscopic finding in the sputum of asthmatics which are spiral

shaped mucus plugs from subepithelial mucuos gland ducts or
bronchioles. These may occur in several different lung diseases
Dittrich plugs (Traube plugs)
Minute, dirty grayish, foul smelling masses of bacteria and fatty acid
crystals in the sputum seen in pulmonary gangrene and fetid
bronchitis.
8. SYNOVIAL FLUID ANALYSIS Sterile containers for microbiology testing

also called Synovia Plain tubes are used for chemistry and immunology testing
is present in areas of the skeleton where friction could develop
such as joints, bursae and tendon sheaths Bulge test
Bathes and lubricates the joints is used to determine if there is an abnormal amount of fluid
joint fluid resembles an egg white surrounding a joint.
Synoviocytes - a three cells deep layer synovial cells found at the surface of
the synovial membrane surrounding the joints
Two types of Synoviocytes based on their physiologic roles
1. Predominating type is actively phagocytic and synthesises degradative

enzymes.
2. Synthesizes and secretes hyaluronate (mucopolysaccharide)
Classification of Joint Disorders

Joint Diseases like Arthritis are common and laboratory analysis of
SF assists in the diagnosis and classification of these conditions
LABORATORY TESTS for SF:
4 Principal categories in the Classification of SF based on Laboratory Exam A. PHYSICAL EXAM
Volume: up to 4 mL
1. Noninflammatory Color & Clarity: Colorless and Clear
2. Inflammatory Yellow/clear - noninflammatory effusions
3. Septic Yellow/cloudy - inflammatory process
4. Hemorrhagic Red/Brown/Xanthochromia - Hemorrhage
Note that these categories partially overlap, several conditions can
occur in the joint at the same time, and variations in test results can Rice bodies - free floating aggregates of tissue seen in Rheumatoid arthritis.
occur depending on the stage of the disease process. Guide for the Ochronotic shards - debris from metal and plastic joint prosthesis, look like
clinicians in the evaluation and diagnosis of joint disease. ground paper
Specimen Collection Viscosity: Very viscous due to high concentration of polymerised

- a noninvasive procedure to determine the existence hyaluronate
of excess fluid around a joint. STRING TEST - used to evaluate the level of SF thickness. 5 cm
ARTHROCENTESIS - procedure for collecting SF from a joint capsule. long before breaking
Heparinized tube is preferred for cell counts,

decreased viscosity of SF indicates the presence of inflammatory URIC ACID

process (<3 cm) like water. Presence is helpful in diagnosing gout
Normal ranges 6 to 8 mg/dL
Clotting - presence of fibrinogen. Tube with heparin may help avoid UA crystal identification is used
clotting SF UA levels may be performed in the lab.
that do not have light polarizing microscope
Mucin Clot
Also known as Rope Test - is an estimation of the integrity LACTIC ACID
of the hyaluronic acid-protein complex (mucin) Rarely measured in SF but can be helpful in diagnosing septic
Normal SF forms a tight ropy clot upon the addition of arthritis.
acetic acid < 25mg/dL but can be as high as 1,000mg/dL in aseptic arthritis.
Good mucin clot indicates good integrity of the hyaluronate.
Poor much clot one that breaks up easily is associated with LACTIC DEHYDROGENASE/LDH
destruction or dilution of hyaluronate. Increased in RA, infectious arthritis, and gout
Normal = 4-6 cm LDH SF can be elevated while serum levels remain normal
When 2-5% acetic acid is added, normal synovial fluid will
form a clot surrounded by clear fluid RHEUMATOID FACTOR/RF
An antibody in immunoglobulin, seen in RA
B. CHEMICAL EXAMINATION OF SF SF RF is being produced by joint tissue +, Serum RF -
PROTEIN C. MICROSCOPIC EXAM OF SF

contains all protein found in plasm, except various high molecular
weight protein (fibrinogen, beta-2 and alpha-2 macroglobulin) can CELL COUNTS
be absent or present in little amount. Saline uses as diluent for SF with high number of cells.
Normal range = 1 to 3 g/dL Normally RBCs < 2000 cells/uL & WBCs <200cells/uL are present in
Increased levels seen in ankylosing spondylitis, arthritis, SF
arthropathies, that accompany Crohn disease, gout, psoriasis, Reiter > than 2000 WBCs is associated with bacterial arthritis, leukocytosis
syndrome, and ulcerative colitis.
DIFFERENTIAL CELL COUNTS
GLUCOSE Normal SF contains small number of lymphocytes and a few
Normal value = 10 mg/dL |< serum levels. neutrophils
SF glucose levels should be interpreted using Serum glucose levels Other cells seen are plasma cell, eosinophils, & Lupus
Joint disorders that are classified as infectious shoes large decreases erythematosus/LE cells.
in SF glucose Septic Arthritis exhibits a high number of neutrophils

Neutrophils present in the later stages of RA may exhibit inclusions CRYSTALS SEEN IN SYNOVIAL FLUID
the contain immune complexes such as IgG, IgM, complement and Monosodium urate crystals - the needle-shaped crystals
RF demonstrate negative birefringence because they are yellow when
aligned with the compensator.
RAGOCYTES/RA - neutrophils that will appear to have dark cytoplasmic Calcium pyrophosphate dehydrate crystals - the rhomboidal crystal
granules demonstrates positive birefringence because it is blue when aligned
with the compensator filter.
CRYSTALS
One of the most important laboratory tests routinely performed D. MICROBIOLOGIC EXAM FOR SF
on SF is microscopic exam of crystals
Pathognomonic of a specific joint disease, thereby enabling a GRAM STAIN
rapid definitive diagnosis when positive provide immediate useful clinical and diagnostic
Temperature and pH changes affect crystal formation and information
solubility. SF should be maintained at room temp. and examined Bacterial originate from the blood
asap. Includes fungi, viruses and mycobacteria
Sensitivity of GS depends on the organism involved
CULTURE
Done whether Gram stain is positive or not
MOLECULAR TESTS
PCR - currently used to identify difficult to detect microorganism,
such as Borrelia burgdorferi that causes Lyme arthritis and
Mycobacterium tuberculosis which can cause osteoarticular
tuberculosis

9. VAGINAL SECRETIONS vagina, and it has lactic acid which is the major metabolic end
product, that maintains the normal acidic pH.
The most common gynecological complains encountered by health care
It can also produce H2O2 which further enhances the healthy acidic
providers are vaginal discharge, discomfort, and odor. The causative agent
environment of the vagina
for the vaginal conditions is distinctly different, the clinical presentations
can be nonspecific and similar. Determining the causative agent before MICROSCOPIC EXAMINATION
initiating therapy is important, and in some cases treating sexual partners is
also necessary to avoid reinfection. Perfomed as soon as possible on vaginal secretion specimens,
particularly for detection of T. vaginalis an actively motile organism
SPECIMEN COLLECTION AND HANDLING Prepare microscope slides using the vaginal swab. Direct wet mount
and 10% KOH preparation. Gram stain is also prepared.
Collected by a healthcare provider during a pelvic examination
Method of collection and container used is specific for the testing
that is to be performed THREE COMON CAUSES OF VAGINAL SECRETIONS INFECTIONS
A warmed speculum is used to visualized the vaginal fornices and
1. BACTERIAL VAGINOSIS
specimen is collected by swabbing the area required for testing-
Most common cause of vaginal infections in women, w/c
vaginal pool, wall, or cervical os (cervical opening to the uterus)
results not from an exogenous pathogen but from an
Swab used is dependent upon the test to be performed. Polyester-
alteration in the normal indigenous bacterial flora of the
tipped swabs on a plastic shafts should be used for bacterial culture.
vagina.
It must be placed immediately into a properly labeled tube or
Complications have shown increased risk in pregnant
container, containing transport media, to be stored and transported
women for premature labor and delivery and low-birth-
at the correct temperature. Cotton has been toxic to N. gonorrhea,
weight infants.
while wooden shafts have been toxic to Chlamydia trachomatis.
Often asymptomatic, with the only complaint of an
A standard patient identification information, an appropriate
unpleasant smell discharge after sexual contact.
me
status, exposure to STDs and use of vaginal lubricants, creams, Single most reliable indicator is the presence of CLUE CELLS
douches. in the wet mount prep.
Notable feature is that WBC are rare. This luck of an
pH/REACTION increase in WBC suggests that the microbial organisms
involved do not invade the subepithelial tissue. Hence the
Vaginal secretions should be determined using commercial pH
condition called VAGINOSIS instead of vaginitis.
paper before the sample swab is placed into saline
Assists in the differential diagnosis of vaginitis
pH range of 3.8 4.5
>4.5 is associated with bacterial vaginosis, trichomoniasis, and
atropic vaginitis. Lactobacilli seen predominantly in a healthy
The consistency and reliability of these findings have culminated in the white, curd-like discharge
classic diagnostic criteria for bacterial vaginosis, w/c requires the presence pH remains normal (pH 3.8 4.5)
of at least 3 of the following 4 features: KOH preparation exam. Reveals budding yeast and/or
pseudohyphae
a. Presence of Clue cells (sloughed off squamous epithelial cells
covered with numerous small thin, curved gram-variable bacilli.
3. TRICHOMONIASIS
b. Positive amine test
Parasitic gynecologic infection caused by Trichomonas
c. Vaginal pH 4.5
vaginalis is among the most common and affects millions of
d. Homogeneous vaginal discharge
women worldwide.
Sexually transmitted, with humans being as its only known
2. CANDIDIASIS
host
Valvovaginal candidiasis (80-92% caused by Candida
In women, trichomonads primarily reside in the vaginal
albicans) is the second most common cause of vaginitis in
mucosa
women. Most adult women have experienced ate least one
episode of vaginal candidiasis and may have had several In men, they infect the urogenital tract
infections. Occurs in celibate as well as sexually active Infection in women can range from an asymptomatic carrier
women and decreases with age, being less common in to a severe, inflammatory condition.
postmenopausal women. Recu
Other Candida species, particularly Candida (Torulopsis) treated simultaneously because of the fact that approx. 35%
glabrata are appearing with increasing frequency, and it is of asymptomatic male partners are positive for T. vaginalis
postulated that this is happening because of an increase in when tested.
self-diagnosis and treatment w/ over-the-counter Trichomoniasis in men is usually asymptomatic or present
antimycotic agents. as urethritis
Are part of normal Vaginal flora In pregnant women, trichomoniasis is a risk factor for
Clinical conditions that predispose an individual to develop preterm rupture of membranes and for premature labor
candidiasis include and delivery.
Pregnancy 50% of women are asymptomatic, the remaining women
Uncontrolled DM complain of a copious frothy, often very unpleasant
Immunosupression smelling discharge that is yellow to greenish, Experience
HIV Infection soreness of the vulva, external dysuria, and dyspareunia
Common infection in women is usually evident by: (difficult or painful sexual intercourse)
vulvovaginal itching Pelvic exam revels vaginal inflammation, and visually the
soreness exocervix is often described as strawberry-like because of
external dysuria numerous punctuate hemorrhages

Most rapid and economical method for diagnosing TREATMENT FOR VAGINAL SECRETION INFECTIONS
trichomoniasis is a direct wet mount. The skill and expertise
Most successful treatment of bacterial vaginosis is orally
of the microscopist directly affect the results of a wet
administered METRONIDAZOLE. New approach to treatment and
mount method.
recolonization of the vagina is the use of lactobacillus-containing
Elevation of pH (5.0 6.0) of the vaginal secretions, reveals
vaginal suppositories.
numerous clumped WBC, KOH preparation often produces a
Topical antimycotic agents from the family of imidazole derivatives
(+) amine test.
predominate, such as a moconazole, clotrimazole. Oral agents
appear to be equally effective and include fluconazole,
4. ATHROPIC VAGINITIS
ketoconazole. Recurrent Candidiasis, defined as four or more
In perimenopausal and postmenopausal women, the vaginal
episodes a year, is a problem for a minority of women and may
epithelium changes because of the reduction in estrogen
require long-term (6 months) antimycotic suppression therapy.
production.
Treatment for T. vaginalis infection in women and men consists of
These changes include thinning of the vaginal epithelium
metronidazole. Oral therapy is preferred because it ensures that all
and decreased glycogen production. As glycogen production
potential sites (vagina, urethra, periurethral glands, prostate,
in the vagina decreases, so does the presence of lactobacilli
epididymis) that may harbor the organism are treated.
and their metabolic by-products lactic acid. These changes
can lead to the development of atrophic vaginitis, with mild
to moderate conditions being asymptomatic. TABLE SHOWING VAGINAL SECRETION FINDINGS
Rare severe cases of atrophic vaginitis, women complain of
vaginal dryness, soreness, dyspareunia (difficult or painful
sexual intercourse), and spotting.
Pelvic examination reveals a thin, diffusely red vaginal
mucosa with little to no vaginal folding.
Alkaline pH usually 5.0. Wet mount reveals numerous WBC
and small number of RBC. In addition to the usual squamous
epithelial cells, parabasal and to a lesser extent basal cells
may be present. KOH preparation and amine test are
negative.
Parabasal cells are the smallest epithelial cells seen on a
typical vaginal smear

DIFFERENTIAL DIAGNOSIS OF VAGINITIS 10. PREGNANCY TEST
Trichomoniasis Candidiasis BV
Complaints Discharge Inc. Severe pruritis Discharge Inc.
burning PURPOSE OF PREGNANCY TEST
Mild Pruritis Mild Pruritis
Vaginal Thin, purulent White curd-like White, fishy Detect human chorionic gonadotropin (hCG)
Discharge frothy homogenous Present in the tissues, blood, and body fluids whether there is living
Vaginal Punctuated Edema Normal placental (chorionic) tissue
Epithelium hemorrhage erythematic
Demonstrated as early as 2 days after the first missed period
Vaginal pH >5 (5.6-6.5) <4.5 >4.5 (4.7-5.7)
May be performed on blood or urine of a pregnant woman
- - +
Microscopic First morning specimen is preferred because this is the most
Trichomonad Candida Clue cells
Examination WBC many WBC some WBC rare concentrated
IMPORTANCE WHY PREGNANCY TEST IS REQUESTED

WHIFF TEST To determine if a woman is pregnant or not because some needs
Potassium hydroxide amine test medication
Uses 10% KOH To determine if there has been a complete or incomplete abortion
Upon application of 10% Potassium hydroxide, to a vaginal swab To differentiate other pelvic diseases
sample, a FISHY ODOR is released, which can suggest To differentiate pregnancy from other uncommon conditions like
trichomoniasis or bacterial vaginosis. hydatidiform mole and chorioepithelioma
Note: The vaginal discharge of patients with BV has a characteristic fishy HUMAN CHORIONIC GONADOTROPIN
odor due to increased activity of anaerobic species. Addition of KOH will Secreted by the cytophoblast cells
augment this odor. Demonstrated in the urine: 10-14 days after conception
Peak level first two months after the missed period
Slowly decreases to a constant low levels 16th week
Disappear 1st week after parturition/giving birth
The developing placenta begins releasing hCG into blood as early as
6 days after implantation. Some hCG also gets passed in the urine
hCG helps to maintain pregnancy and affects the development of
fetus

Levels of hCG increase steadily in the first 14 to 16 weeks following TWO GROUPS OF TEST FOR PREGNANCY
LMP, peak around the 14th week following LMP, and then decrease
1. BIOASSAY TEST
gradually
The amount that hCG increases early in pregnancy can give utilize the hyperemic or spermatozoa and ova producing effect of
information about pregnancy and the health of the baby. Shortly hCG on animal gonads.
after delivery, hCG can no longer be found in the blood Test animals used are rabbits, mice, rats, frogs, toads.
More hCG is released in a multiple pregnancy, such as twins or
2. IMMUNOLOGIC TEST
triplets, than in a single pregnancy
Less hCG is released if the fertilized egg implants in a place other Agglutination -inhibition and hemagglutination principle
than the uterus, such as in a fallopian tube. This is called an ectopic using red cells or latex particles sensitized by hCG
pregnancy
METHODS
VARIATIONS IN THE hCG LEVEL
A. Physician Tests
Increase in multiple pregnancies, H-mole, chorioepithelioma,
B. Home Pregnancy Tests
Malignant teratomas.
Decrease in incomplete abortion C. Test based on detection of hCG
Pregnancy Test should be negative in 3-4 days after delivery, within
Human Chorionic Gonadotropin
1 week. Should become negative after delivery of a mole
Positive Test persistent mole, ectopic pregnancy (+until the death of a hormone secreted by the placenta; it is the substance detected in
the placenta) pregnancy tests
POSITIVE PREGNANCY TEST IN CASES OF: glycoprotein hormone in increased amounts in pregnant women and
in some tumors
Seminoma hydatidiform mole: rare mass that forms inside the uterus at the
Choriocarcinoma beginning of a pregnancy
Germ cell tumors Germ cell tumors in both men and women
Hydatidiform mole formation
D. To confirm pregnancy - blood or urine test
Teratoma with elements of choriocarcinoma
Islet cell tumor BLOOD TEST - 3 days after conception
URINE TEST - 2 weeks after missed period

OBSTETRICIAN - OBGYN Latex Agglutination Inhibition Tests (Urine)
Doctor specializes in pregnancy Radioreceptor Assay

Examination includes:
Blood pressure, pulse, respiration, weight
Pelvic measurement NOTE: High Sensitivity pregnancy tests (<1mIU/mL) - sometimes false positive
Analysis of urine - condition of kidneys becaause of endogenous pituitary hCG
Blood Test - check for anemia
History of illness and operations
Pregnancy Test application of ELISA assay. In home pregnancy test,
IMMUNOLOGIC PREGNANCY TEST
direct measurement of antigens, such as human chorionic
Rapid, sensitive and accurate gonadotropin (hCG)
Eliminate the use of animals Home pregnancy test utilizes the principle of sandwich enzyme
immunoassay, with a unique mono-antibody combination specific
ASSAYS:
against hCG present in urine/serum
Radioimmunoassay (RIA)
AGGLUTINATION-INHIBITION TEST
Sensitivity: 5 mIU/mL
Urine from pregnant woman +hCG is added to the reagent
Quantitative
hCG neutralizes the Ab which fails to act on the Ag. AGG does not
can determine gestational age
occur (+)
uses serum
Homogeneous settling of the non-agglutinated Ag
Immunoradiometric Assay Negative shows agglutination
Sensitivity: 150mIU/mL
EVALUATION OF PLACENTAL FUNCTION IN ESTABLISHED PREGNANCY
ELISA
THREE METHODS:
Sensitivity: 25-150 mIU/mL
Qualitative 1. Amniotic fluid in Rh sensitization
Anti-hCG bound to membrane
2. Assay of Urinary Estriol Excretion
Can detect hCG 7-10days after conception
uses serum, urine PLACENTAL ESTRIOL ASSAY
Fluoroimmunoasay Methods: Gas Chromatography
Sensitivity: 1mIU/mL
Normal Pregnancy = urinary estrogen excretion rises constantly until term

that it rapidly falls
Placental Insufficiency: A drop of over 70% of the preceding level during

pregnancy
3. Assay of heat-stable serum ALP
Heat-Stable ALP
Placenta - major source of heat-stable isoenzyme of ALP
Total serum ALP value. Serum is inactivated at 56*C for 30mins and
assayed
Inadequate placental function- progressive fall of serial values

Microscopic Exam Body Fluids

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CLINICAL MICROSCOPY: Microscopic and Body Fluids MLS INTERNS 2019 NOTES TEAM

Renal Tubular Epithelial cells

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2. EXPECTED STAINED COLOR WHEN THE FOLLOWING FORMED ELEMENTS

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DESCRIBE HOW THE LABORATORY CAN DEMONSTRATE FAT/LIPIDS IN

Gram stain is used in microbiology to differentiate bacteria into gram

neutrophils. Eosinophils are increased in acute interstitial nephritis,

cytoplasmic granules, and absence of disintegration

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TRANSITIONAL EPITHELIAL CELLS IN URINE CLINICAL SIGNIFICANCE OF TRANSITIONAL CELLS

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CLASSIFICATION OF CASTS Staining with Sternheimer-Malbin stains enhances their visualization.

These casts form whenever any disorder damages the glomerulus,

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Note: Performing a Gram Stain may facilitate

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o Entamoeba histolytica gram-stain technique may be helpful in visualizing these

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HIPPURIC ACID CRYSTALS AND THEIR SIGNIFICANCE

SULFONAMIDE CRYSTALS AND THEIR SIGNIFICANCE D. Acetylsulfadiazine

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THE FOLLOWING FORMS THE AF:

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FETAL DISTRESS TESTING

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2. FOAM STABILITY INDEX/ FOAM TEST/ SHAKE TEST CULTURE

Test not affected by hemolyzed blood or meconium

5. OPTICAL DENSITY AT 650 NM

6. MICROVISCOSITY FLUORESCENCE POLARIZATION ASSAY

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2. CEREBRO SPINAL FLUID (CSF) PURPOSE OF CSF EXAMINATION

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2. COLOR CHEMICAL EXAMINATION

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2. ALBUMIN & IgG 2. CSF GLUCOSE

NORMAL BACTERIAL VIRAL FUNGAL or

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METHODS OF CELL COUNT

USING AN IMPROVED NEUBAUER CHAMBER GROSS APPEARANCE

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DIFFERENTIAL COUNT TABLE SHOWING DIFFERENT CONDITIONS IN CSF

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Gullain- 0-100 L N >100 N FLUOROCHROME SMEAR to detect M. tuberculosis in CSF.

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5. SEROLOGIC EXAMINATION 7. GRAM STAIN

6. LIMULUS AMEBOCYTE LYSATE

POLYMERASE CHAIN REACTION (PCR) is a technique used to

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1. HYDROCHLORIC ACID - secreted by parietal cells hydrolyzes peptides

DISORDERS OF GASTROINTESTINAL TRACT

A. UPPER GASTROINTESTINAL BLEEDING (UGIB)

bleeding in the esophagus, stomach, or duodenum,

Repeated hemoglobin levels and occult blood

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common disorder of the GI tract Collection of the Specimen

Due to increased secretions Two Types of Gastric Juice Collection

common cancer of the GI tract