GMT 109: Genitourinary System

Course

PHASE I - M.D.
(2022/2023)

URINALYSIS (PRACTICAL)
Prepared by

Assoc. Prof. Dr. Julia Omar

Prof. Dr. K.N.S.Sirajudeen
Dr. Noorazliyana Shafii
Dr. Aniza Mohd. Jelani
Dr. Zulkarnain Mustapha
Dr Wan Norlina Wan Azman
Dr. Tuan Salwani Tuan Ismail
Dr. Noor Azlin Azraini Yusof @ Che Soh

Department of Chemical Pathology

School of Medical Sciences
Universiti Sains Malaysia
Kubang Kerian, Kelantan
Malaysia
 Contents

Page

1. Objective 2

2. Introduction 2

3. Collection Of Urine Specimen 3

4. Physical Examination of Urine 5

5. Chemical Analysis 6

6. Microscopic Examination 9

7. Reference Values 10

8. Practical (hands on) 16

Genitourinary system (GMT 109): Urinalysis (Practical) 1

 URINALYSIS

1. OBJECTIVES

1. To understand what is urinalysis (Urine FEME - Full Examination

Microscopic Examination) and its components
2. To perform chemical analysis for the presence of protein using
reagent strips
3. To perform microscopic examination of urine

2. INTRODUCTION

Urine is continuously formed by the kidneys. It is an ultrafiltrate of plasma from

which glucose, amino acids, water and other substances essential to body
metabolism have been reabsorbed. Two important characteristics of a urine
specimen make it an integral part of the patient examination.

Urine
a) It is readily available and easily collected
b) It contains information about many of the body’s major metabolic functions
and it can be obtained by simple lab tests.

Examination of urine analysis includes physical examination (macroscopic

examination), chemical analysis (reagent strips / dipstick testing) and
examination of urine sediments (microscopic examination).

Physical Urine sediments

(macroscopic examination) Chemical analysis (microscopic examination)

Volume Specific gravity (SG) RBC

Colour & Turbidity pH WBC
Odour Protein Crystals
Glucose Casts
Ketones Epithelial cells
Bilirubin Miscellaneous
Urobilinogen (Bacteria, Yeast, Sperm etc)
Hemoglobin
Nitrites

Genitourinary system (GMT 109): Urinalysis (Practical) 2

 3. COLLECTION OF URINE SPECIMEN

Urine is biologically hazardous material and universal precautions should be

taken while handling the specimen.

Specimen container:

 Specimens must be collected in a clean and dry container

 Disposable containers are recommended
 Should be available in various sizes, shapes and plastic bags with
adhesives for pediatric specimen

Methods of urine specimen collection / types of specimen

1. Random collection
Taken at any time of day with no precautions regarding contamination.
The sample may be dilute, isotonic, or hypertonic and may contain white
cells, bacteria and squamous epithelium as contaminants. In females, the
specimen may contain vaginal contaminants such as trichomonads, yeast
and red cells.

Purpose: Routine screening

2. Early morning collection

The sample is taken before ingestion of any fluid. This is usually
hypertonic and reflects the ability of the kidney to concentrate urine during
dehydration which occurs overnight. If all fluid ingestion has been avoided
since 6 p.m. the previous day, the specific gravity usually exceeds 1.022
in healthy individuals.

Purpose: Routine screening

Pregnancy tests

3. Clean-catch, midstream urine specimen

Collected after cleansing the external urethral meatus. A cotton sponge
soaked with benzalkonium hydrochloride is useful and non-irritating for
this purpose. A midstream urine is one in which the first half of the bladder
urine is discarded and the collection vessel is introduced into the urinary
stream to catch the last half. The first half of the stream serves to flush
contaminating cells and microbes from the outer urethra prior to collection.

Purpose: Routine screening

Bacterial culture

Genitourinary system (GMT 109): Urinalysis (Practical) 3

 4. Catheterization of the bladder
Through the urethra for urine collection is carried out only in special
circumstances, eg. in a comatose or confused patient. The risk of this
procedure is introducing infection and traumatizing the urethra and
bladder, thus producing iatrogenic infection or hematuria.

Purpose: Bacterial culture

5. Suprapubic transabdominal needle aspiration of the bladder

When done under ideal conditions, this provides the purest sampling of
bladder urine. This is a good method for infants and small children.

Purpose: Bacterial culture and cytology

Specimen delivery / preservation:

Specimen must be delivered to the laboratory promptly and tested within 1 hour.
If not it should be refrigerated or added with appropriate chemical preservative.

Changes in unpreserved urine (at room temperature for more than 1 hour)

 Increased pH from the breakdown of urea to ammonia by urease

producing bacteria
 Decreased glucose due to glycolysis and bacterial utilization
 Decreased ketones because of volatilization
 Decreased bilirubin from exposure to light
 Deceased urobilinogen
 Increased nitrite due to bacterial reduction of nitrate
 Increased bacteria
 Increased turbidity caused by bacterial growth and possible precipitation of
amorphous material
 Disintegration of red blood cells & casts especially in dil. alkaline urine
 Changes in colour due to oxidation or reduction of metabolites

Genitourinary system (GMT 109): Urinalysis (Practical) 4

 4. PHYSICAL EXAMINATION OF THE URINE

Volume: Normal urine volume is 750-2000 ml/24 hr

Colour:

 First part of the urinalysis is direct visual observation.

 Normal, fresh urine is clear, pale to dark yellow or amber in colour and is
due to normal urine pigments.
 Colour of urine may change in many disease states because of the
presence pigments eg. Bilirubin – dark coloured / tea coloured urine
 A red or red-brown color could be due to food dye, eating fresh beets,
drugs or presence of either hemoglobin or myoglobin.

Turbidity:

 Turbidity or cloudiness may be caused by excessive cellular materials or

protein in urine (or) may develop from crystallization (or) precipitation of
salts upon standing at room temperature (or) in the refrigerator.
 Clearing of the specimen after adding a small amount of acid indicates
precipitation of salts for the probable cause of turbidity.

Odour:

 Normal freshly voided urine gives very little smell and is due to the
presence of volatile acids
 Prolonged long standing urine – ammonia like odour is due to bacterial
decomposition of urea
 Diabetic urine – Fruity odour due to increased excretion of acetone
 Urine of patients with urinary tract infection – Foul-smell
 Certain food/drugs may cause a characteristic odour

Genitourinary system (GMT 109): Urinalysis (Practical) 5

 5. CHEMICAL ANALYSIS (To perform)

Reagent (dipstick) strip testing

Method

1. Understand the principle and significance of the test

2. Mix the specimen well
3. Dip briefly and completely into specimen
4. Remove excess urine when withdrawing strip from specimen
5. Compare reaction colours with manufacturer’s chart under a good light
source after 30-60 seconds.
6. Be alert for the presence of interfering substances

1. Protein

Dipsticks detect protein by production of color with an indicator dye,

Bromphenol blue.

Dipstick screening for protein is done on whole urine.

However semi-quantitative tests for urine protein should be performed on

the supernatant of centrifuged urine since the cells suspended in normal
urine can produce a falsely high estimation of protein.

Normally, plasma proteins filtered at the glomerulus are reabsorbed by the

renal tubules. However a small amount of filtered plasma proteins and
protein secreted by the nephrons can be found in normal urine.

Normal total protein excretion does not usually exceed 150 mg/24 hours.
More than 150 mg/day is defined as PROTEINURIA (protein in urine)

Proteinuria > 3.5 gm/24 hours is severe and found in cases of Nephrotic
Syndrome.

Fever, exposure to heat or cold, excessive exercise and emotional stress

may cause proteinuria and usually such findings are temporary.

Excessive and regular excretion of proteins (pathological proteinuria)

usually suggests renal disease and results from damage to the glomerulus

Genitourinary system (GMT 109): Urinalysis (Practical) 6

 such as glomerulonephritis or defect in the reabsorption process of the
renal tubules such as pyelonephritis, tubular acidosis etc

Proteinuria with symptoms of increase frequency and painful micturition is

usually suggestive of Urinary Tract Infection

2. Specific gravity (SG)

Specific gravity (which is directly proportional to urine osmolality)

measures urine density or the ability of the kidney to concentrate or
dilute the urine over that of plasma.

Dipsticks are available for the measurement of specific gravity in

approximations. Specific gravity between 1.002 and 1.035 on a random
sample considered normal.

Reduced specific gravity as a consequence of reduced concentrating

ability of the kidney is found in cases such as Diabetes insipidus,
glomerulonephritis, pyelonephritis and other renal disorders.

Elevated specific gravity due to excessive loss of water is found in cases

such as fever, vomiting, diarrhoea, adrenal insufficiency, liver diseases &
congestive heart failure.

.
3. pH

The glomerular filtrate of blood plasma is usually acidified by renal tubules

and collecting ducts from a pH of 7.4 to about 6 in the final urine.
However, depending on the acid-base status, urinary pH may range from
as low as 4.5 to as high as 8.0. The change to the acid side of 7.4 is
accomplished in the distal convoluted tubule and the collecting duct.

4. Glucose

Dipsticks employing the glucose oxidase reaction for screening are

specific for glucose. Other reducing sugars such as galactose and
fructose are screened by other methods such as modified Benedict's
copper reduction test.

Less than 0.1% of glucose normally filtered by the glomerulus appears in

urine (< 130 mg/24hr).

Genitourinary system (GMT 109): Urinalysis (Practical) 7

 GLYCOSURIA (excess sugar in urine) generally occurs whenever the
blood glucose level exceeds the reabsorption capacity of the renal
tubules (renal threshold).

5. Ketones

Ketones (acetone, acetoacetic acid, beta-hydroxybutyric acid) resulting

from either diabetic ketoacidosis or some other form of calorie
deprivation (starvation) are easily detected using either dipsticks or test
tablets containing sodium nitroprusside.

6. Bilirubin

BILIRUBINURIA (bilirubin in urine) indicates the presence of

hepatocellular disease or intra- or extrahepatic obstruction. Excretion of
bilirubin in the urine will reach significant levels in any disease process
that increases the amount of conjugated-bilirubin and not due to the
increases of unconjugated-bilirubin.

7. Urobilinogen

Bacterial action in the intestinal tract converts the bilirubin to urobilinogen

and estimated half of the urobilinogen is reabsorbed into the portal
circulation and re-excreted by the liver and small amounts are normally
excreted in the urine.

Urobilinogen excretion in the urine is increased by any conditions that

causes an increase in the production of bilirubin and is decreased if there
is obstruction to the bile flow which affects the transport of conjugated
bilirubin to intestine.

8. Hemoglobin

HEMOGLOBINURIA (hemoglobin in urine) may indicate hematologic

disorders, severe infectious diseases, chemicals poisoning or a neoplastic
disease.

Genitourinary system (GMT 109): Urinalysis (Practical) 8

 9. Nitrite

A positive nitrite test indicates that bacteria may be present in significant

numbers in urine. Gram negative rods such as E. coli are more likely to
give a positive result.

6. MICROSCOPIC EXAMINATION (To perform)

Method

1. A sample of well-mixed urine (usually 10-15 ml) is centrifuged in a test

tube at relatively low speed (about 2-3,000 rpm) for 5-10 minutes (until a
moderately cohesive button is produced at the bottom of the tube).
2. The supernate is decanted to a volume of 0.2 to 0.5 ml.
3. The sediment is resuspended in the remaining supernate by flicking the
bottom of the tube several times.
4. A drop of resuspended sediment is poured onto a glass slide and covered
with a coverslip.
5. Examine under low power field (LPF) to identify crystals, casts, squamous
cells and other large objects. The numbers of casts seen are usually
reported as number of each type found per low power field (LPF) example
5-10 hyaline casts/LPF.
6. Next, examine under high power field to identify the type of crystals, cells,
and bacteria. The various types of cells are usually described as the
number of each type found per average high power field (HPF) example:
1-5 WBC/HPF.

Genitourinary system (GMT 109): Urinalysis (Practical) 9

7. REFERENCE VALUES

Components Reference Value

RBCs 0-3 / hpf (male), 0-5 / hpf (female)

WBCs 0-4 / hpf

Epith/Hpf Occasional (may be higher in females)

Casts/Lpf Occasional hyaline

Bacteria Negative

Crystals Types present vary with pH (Crystals such as cystine, leucine,

tyrosine and cholesterol are considered abnormal)
# Hpf – high power field Lpf – low power field

Components

1. Red blood cells

Biconcave discoid shape

Central pallor
Anucleated

Hematuria (blood in urine) is the presence of abnormal numbers of red

cells in urine. The causes could be due to glomerular damage, tumors
which erode the urinary tract anywhere along its length, kidney trauma,
urinary tract stones, renal infarcts, acute tubular necrosis, upper and
lower urinary tract infections, nephrotoxins and physical stress.

Red cells may also contaminate the urine from the vagina in menstruating
women or from trauma produced by bladder catheterization.

Theoretically, no red cells should be found however some are found in the
urine of healthy individuals. The RBC's may appear normally shaped

Genitourinary system (GMT 109): Urinalysis (Practical) 10

 (biconcave), swollen by dilute urine or crenated by concentrated urine.
Both swollen, partly hemolyzed and crenated RBC's are sometimes
difficult to distinguish from WBC's

The presence of dysmorphic RBC's (odd shapes) in urine as a

consequence of being distorted via passage suggests a glomerular
disease such as a glomerulonephritis.

2. White blood cells

Lobed nuclei
Granular cytoplasm

Pyuria refers to the presence of abnormal numbers of leukocytes that may

appear with urinary tract infection. Usually, the WBC's are granulocytes.

If two or more leukocytes per each high power field appear in non-
contaminated urine, the specimen is probably abnormal. Leukocytes have
lobed nuclei and granular cytoplasm.

3. Epithelial cells

Epithelial cells Squamous epithelial cells

Genitourinary system (GMT 109): Urinalysis (Practical) 11

 Renal tubular epithelial cells are usually larger than granulocytes. It
contains large round or oval nucleus and normally sloughs into the urine in
small numbers. The number sloughed is increased in conditions such as
nephrotic syndrome and conditions leading to tubular degeneration.

Transitional epithelial cells from the renal pelvis, ureter, or bladder have
more regular cell borders, larger nuclei, and smaller overall size than
squamous epithelium.

Squamous epithelial cells in the urine are usually from the skin surface or
from the outer urethra. They represent possibility of contamination of the
specimen with skin flora.

4. Crystals

Common crystals seen even in healthy patients include calcium oxalate,

triple phosphate crystals and amorphous phosphates. Uric acid crystals
are seen in cases of hyperuricemia. Other crystals are uncommon.

Octahedral shaped
Tiny, colourless or dark granules
Can be either isolated or in clumps.

Dumbbell shaped

Calcium oxalate crystals Amorphous phosphates crystals

(Picture adopted from eClinPath)

Genitourinary system (GMT 109): Urinalysis (Practical) 12

 Three dimensional, prism-like crystals Rhomboid shape

Triple phosphate crystals Uric acid crystals

5. Urinary Casts

Urinary casts are formed only in the distal convoluted tubule (DCT) or the
collecting duct. The proximal convoluted tubule (PCT) and loop of Henle
are not locations for cast formation.

Type of casts Explanation

Hyaline casts
- Are composed primarily of a
mucoprotein (Tamm-Horsfall protein)
secreted by tubule cells in the
collecting duct.
- The factors which favor protein cast
formation are low flow rate, high salt
concentration and low pH, all of which
favor protein denaturation and
precipitation.
- Hyaline casts can be seen in healthy
patients.

Protein casts
- Protein casts with long, thin tails
formed at the junction of Henle's loop
and the distal convoluted tubule are
called cylindroids.

Genitourinary system (GMT 109): Urinalysis (Practical) 13

 Red blood cell casts
- Are indicative of glomerulonephritis,
with leakage of RBC's from glomeruli
or severe tubular damage

(Picture adopted from Urine Sediment Examination in the

Diagnosis and Management of Kidney Disease: Core
Curriculum 2019 Am J Kidney Dis. 73(2): 258-272)

White blood cell casts

- Are most typical for acute
pyelonephritis and glomerulonephritis.
- Their presence indicates inflammation
of the kidney.

(Picture adopted from www.nephsim.com)

Genitourinary system (GMT 109): Urinalysis (Practical) 14

 Cellular casts
- Are casts that remain in the nephron
for some time, before they are flushed
into the urinary bladder.
- The cells may degenerate to become a
coarsely granular cast later to finely
granular cast and ultimately to waxy
cast.
- Granular and waxy casts are derived
from renal tubular cell casts and broad
casts are from damaged and dilated
tubules which are seen in end-stage
chronic renal disease.

6. Miscellaneous

Budding yeast
Bacteria

Bacteria are common in urine specimens because of the abundant normal

microbial flora of the vagina or external urethral meatus.

Diagnosis of bacteriuria in a case of suspected urinary tract infection

requires culture. Multiple organisms reflect contamination. However, the
presence of any organism in catheterized or suprapubic tap specimens
should be considered significant.

Yeast cells may be contaminants or represent a true yeast infection. They

are distinguished by their tendency to bud.

Genitourinary system (GMT 109): Urinalysis (Practical) 15

8. PRACTICAL: URINALYSIS (HANDS ON)

A 35-year-old female was seen at the Outpatient Clinic with the complaints of
painful micturition. Random urine sample was taken for examination.

Perform the urinalysis and state your findings

1. CHEMICAL ANALYSIS using reagent strips

Urine Protein: ________________________

2. MICROSCOPIC EXAMINATION

Components Findings

RBC

WBC

Epithelial cells

Casts

Bacteria

Crystals

3. State the diagnosis.

Genitourinary system (GMT 109): Urinalysis (Practical) 16