MEDT 25: VIRTUAL INTERNSHIP
HISTOPATHOLOGY SECTION
ACTIVITY # 1: Tissue Processing, Staining and Troubleshooting
Name DOMINGO, JAME-ANN S.
:
Instruction: Analyze each case carefully and answer the given questions. Back-up your
answers by providing reliable references. Use APA 7th edition.
Case
Patient A was admitted to the hospital due to
body weakness, edema, chest pains and
extreme weight loss. After mammogram and
other series of tests was done, her familial
history was also taken. Her family had history
of carcinoma with her aunt dying of breast
cancer. After much consideration, it was
concluded that patient had stage 2 breast
cancer. IV meds were started and patient
was advised to undergo surgery to remove
her breast tissue due to the massive tumor
seen consuming the normal cells. Patient
gives her consent for operation. Left breast
tissue along with the lymph nodes were sent
to the lab for histopathologic examinations.
What type of incisions should be made and
subjected for embedding and microtomy and
what should pathologist look closely when
histotechnicians process the specimen
given?
Date: October 07, 2022
Answer
Based on the situation of the patient,
modified radical mastectomy should be done.
Modified radical mastectomy is the primary
method of treatment for breast cancer. It
involves removing the entire breast, including
the skin, areola, nipple, and most axillary lymph
nodes, but leaving the pectoralis major muscle
intact.
During specimen processing, the
following are some of the things that pathologist
should look closely;
 Whether the tumor is invasive: It is
critical for the pathologist to note how much
the tumor has grown into nearby healthy
tissue in the case of invasive tumors.
 Tumor Grade: The grade describes how
cancer cells appear in comparison to
healthy cells. In general, the pathologist
looks for differences in cell size, shape,
and staining characteristics.
 Mitotic Rate: The pathologist should note
how many cells are dividing.
 Tumor Margin: Another important
consideration is the presence of cancer
cells at the biopsy sample's margins, or
edges. A "positive" or "involved" margin
indicates the presence of cancer cells. This
implies that cancerous cells are still
present in the body.
 Lymph Nodes: The pathologist will also
examine the cancer to see if it has spread
to nearby lymph nodes or other organs.

Serous fluids like pericardial, peritoneal and
pleural fluid should be sent immediately to
the laboratory for cytology processing to
ensure integrity of cells were well-preserved.
In cases where specimen was at the
operating room and was sent to the
laboratory two hours after collection, what
should be the next mode of action if you were
the medical technologist? What physical
changes would you observe in that specimen
and why do you think it happens?
Reference(s):
American Society of Clinical Oncology. (2019).
Reading A Pathology Report. Retrieved from
https://www.cancer.net/navigating-cancer-
care/diagnosing-cancer/reports-and-
results/reading-pathology-
report#:~:text=Grade%20describes%20how%20
the%20cancer,staining%20features%20of%20th
e%20cells.
Kuwajerwala, N. K. (2021). Modified Radical
Mastectomy. Retrieved from
https://emedicine.medscape.com/article/183010
5-
overview#:~:text=A%20modified%20radical%20
mastectomy%20is,of%20treatment%20for%20br
east%20cancer.
Fluid specimens should ideally be sent to
the laboratory immediately and processed
within 2 hours. Except for microbiological
cultures, if a delay is expected, the sample
should be kept at 4 degree Celsius until
analysis. Although the cytomorphological
features of refrigerated samples are well
preserved for at least 72 hours, a delay of more
than 48 hours is unacceptable.
Delays in processing may result in a
degenerating smear image with loss of cell
morphology and a high concentration of
bacteria in the smear background. Air-drying
artifacts such as pale stained nuclei, a lack of
differential cytoplasmic staining, and
cytoplasmic and nuclear eosinophilia can result
from a delay in fixation.
Reference(s):
Porcel, J. M. (n.d.). Handling Pleural Fluid
Samples for Routine Analyses. Retrieved from
https://toraks.org.tr/site/sf/books/pre_migration/
5
406770df55f10e9ea909fa2dd50affa793710a28
b
e6182251502e924f8d24f4.pdf
National Cancer Control Programme. (2005).
Manuals for Training in Cancer Control.
Retrieved from
https://screening.iarc.fr/doc/Cancer_resource_
M
\anual_3_Cytology_New.pdf
Senile male patients often complain difficulty
in urinating producing minute outputs but still
feeling their bladder full. In such cases where
hyperplasia or cancer is considered, TURP is
advised as a medical procedure to check the
reason for urinary obstruction. Upon
receiving the specimen for processing, you
receive a small container filled with prostatic
chips. Do you consider it a small, large or
radical specimen? Why? What process in the
histopathology is utilized in handling such
kinds of specimen and how do you gross it?
How does benign prostate hyperplasia
contribute to urinary obstruction? How about
prostate cancer?
Transurethral resection of the prostate
(TURP) is an endoscopic procedure used to
remove the prostate. The lack of description of
the prostatic chips makes it hard to consider if it
is small, large, or radical specimen. In
processing these kinds of specimen accurate
patient identification, and adequate fixation
must be done. After that, sample receiving and
entering the specimen details should be
completed. In grossing the specimen, the
following should be followed:
 Weigh and measure tissue aggregate in 3D
 Specimens weighing less than or equal to
12 gm: Submit entirely in mesh bags
(should fit in approximately 12 cassettes).
 Specimens weighing more than 12 gm:
Submit 12 grams in mesh bags (in
approximately 12 cassettes), and an
additional cassette per each additional 5
grams.
Benign prostatic hyperplasia (BPH) is a
benign adenomatous overgrowth of the
periurethral prostate gland. Symptoms of
bladder outlet obstruction include a weak
stream, hesitancy, urinary frequency, urgency,
nocturia, incomplete emptying, terminal
dribbling, overflow or urge incontinence, and
complete urinary retention.
Urine outflow becomes increasingly
obstructed as the lumen of the prostatic urethra
narrows and lengthens. Increased pressure
caused by micturition and bladder distention
can lead to bladder detrusor hypertrophy,
trabeculation, cellule formation, and diverticula.
Incomplete bladder emptying causes stasis and
increases the likelihood of calculus formation
and infection. Even if the obstruction is only
partial, prolonged urinary tract obstruction can
cause hydronephrosis and impair renal
function.
BPH is an abbreviation for benign prostatic
hyperplasia. Benign refers to "not cancerous,"
and hyperplasia refers to abnormal cell growth.
As a result, the prostate grows in size. BPH is
not associated with cancer and does not
increase your risk of developing it; however, the
symptoms of BPH and prostate cancer can be
similar.
 Reference(s):

Andriole, G. (2022). Benign Prostatic Hyperplasia

(BPH). Retrieved from
https://www.merckmanuals.com/professional/genitourinar
y-disorders/benign-prostate-disease/benign-prostatic-
hyperplasia-bph

Baradhi, K. & Ng, M. (2022). Benign Prostatic

Hyperplasia. Retrieved from
https://www.ncbi.nlm.nih.gov/books/NBK55892
0/#:~:text=The%20development%20of%20beni
gn%20prostatic,in%20clinical%20manifestation
s%20of%20lower

National Cancer Institute. (n.d.). Understanding

Prostate Changes: A Health Guide for Men.
Retrieved from
https://www.cancer.gov/types/prostate/understa
nding-prostate-changes

Chargui, S. & Stormont, G. (2021).

Transurethral Resection Of The Prostate.
Retrieved from
https://www.ncbi.nlm.nih.gov/books/NBK56088
4/

Gross Pathology Manual. (n.d.). Prostate:

TURP. Retrieved from
https://voices.uchicago.edu/grosspathology/gu-
renal/prostate-turp/
A medical technologist sometimes
accompanies the pathologist when it an order
for UTZ or CT-guided biopsy is done. A
radiologist diagnostically images the target
site for collection and inserts the needle to
suction portions of the tissue or mass. In the
case of pathologist, s/he is there to check if
the sample is enough for processing or
another collection is needed. A medical
technologist smears, stains and mounts the
slides for the pathologist to observe them
under the microscope. In cases of biopsy
where pull apart is the ideal method, how
many slides will you generate if the
radiologist places the specimen in four of the
slides you prepare? How will you fix and stain
the slides? And how do you perform cell
block?
Pull apart method is accomplished by
applying a drop of secretion or sediment to one
side of a clean slide. To start the flow of
materials, the material must be evenly
distributed across the surface of the two slides
in opposite directions. The two slides are then
separated in a single continuous motion, and
the specimen is placed under the microscope
for immediate examination, or vital stain is
applied.This is useful for preparing smears of
thick secretions such as serous fluids,
concentrated sputum, gastro-intestinal lavage
samples, and blood smears.
If the radiologist places the specimen in 4
slides, then 8 slides will be utilized using this
method. After placing the specimen on the
slides, both slides were immersed in 95%
ethanol and were stained by hematoxylin and
eosin (H and E).
Following smearing preparations, the
needles and syringes used to obtain fine-
needle aspirates were rinsed in a specimen
container with 10 mL of 50% ethanol. Any
remaining clots or tissue in the needle hubs
were carefully removed in the laboratory using
another needle and rinsed in 50% ethanol. The
entire material was centrifuged at 4,000 rpm for
6 minutes in a 10-mL disposable centrifuge
tube to produce one or more cell pellets (1
pellet in most cases).
If the centrifuged deposits were too thick,
the material was divided into several tubes for
multiple cell blocks before being fixed in NAFS
solution. After 45 minutes of fixation, the fixed
cell pellets were recentrifuged at 4,000 rpm for
6 minutes. Following centrifugation, these
pellets should detach or be easily removed with
a disposable Pasteur pipette.
The cell pellets were wrapped in crayon
paper, placed in a cassette, and stored in 80%
ethanol until ready for processing in the
automatic tissue processor using the following
13-hour processing schedule: 80% ethanol with
1 change (2.5 hours), 95% ethanol (1 hour),
100% ethanol, 4 times (1 hour each), 1:1
ethanol/xylene (1 hour), xylene, 3 times (1 hour
each), paraffin wax, 60°C (1 hour), vacuum
impregnation at 20 lb. for half an hour The
paraffin-embedded cell blocks were sectioned
at 3 m thickness.
Reference(s):

Lecture Notes in Medical Technology. (n.d.).

Examination of Tissues. Retrieved from
http://mt-
lectures.blogspot.com/2017/10/examination-of-
tissues.html

Hahn, et al. (n.d.). Cytopathologic Touch

Preparations (Imprints) from Core Needle
Biopsies: Accuracy Compared with That of Fine
Needle Aspirate. Retrieved from
https://www.ajronline.org/doi/pdf/10.2214/ajr.16
5.5.7572518

Horn, M., Narayan, E., Nathan, N., & Smith, M.

(n.d.). Cell Block Cytology. Retrieved from
https://watermark.silverchair.com/ajcpath114-
0599.pdf?token=AQECAHi208BE49Ooan9kkh
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When a woman has given birth or had sexual
intercourse, an OB advises for pap smear
yearly to check for any presence of
abnormality in the reproductive area. In
cases were suspected bacterial vaginosis is
considered, what is the pH of the vagina that
promotes the infection? What cells are seen
that might conclude such infection? Describe
the characteristics of normal cells vs cells
with bacterial vaginosis in pap smear?
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Bacterial vaginosis occurs when there is a high
concentration of Gardnerella bacteria and a low
concentration of Lactobacillus bacteria, causing
pH levels to decrease. A normal vaginal PH is
around 4 (acidic), whereas with bacterial
vaginosis, vaginal pH is at 7 or higher (less
acidic).
The presence of clue cells in the vaginal
fluid indicates that a women has a bacterial
vaginosis. In vaginal smears from women with
bacterial vaginosis, epithelial cells were
covered by adherent gram-negative rods. The
gram-negative bacteria adhering to the clue
cells were identified using immunofluorescence
studies. Long-term, multiple, small-inoculum
immunization of rabbits yielded specific antisera
to four common gram-negative vaginal bacteria
(Gardnerella, Bacteroides, Fusobacterium, and
Mobiluncus). Absorption against whole cells of
heterologous bacteria and serial dilution were
used to remove cross-reactivity with
heterologous common vaginal bacteria.
Gardnerella vaginalis was found adhering to the
surface of clue cells and on the surface of
exfoliated vaginal epithelial cells significantly
more frequently and in greater numbers than
Mobiluncus, Bacteroides, and Fusobacterium,
indicating that this gram-negative bacteria is
responsible for clue cell formation.
Vaginal discharge is normally white,
nonhomogeneous, and viscous. It contains
vaginal squamous epithelial cells in a serous
transudate, as well as sebaceous, sweat, and
Bartholin's gland material, and cervix
secretions. A small number of
polymorphonuclear leukocytes, most likely from
the cervix, may be seen. The pH is less than
4.5, typically ranging between 3.8 and 4.2.
Lactobacilli, large gram-positive rods, are the
dominant organisms. But in a vaginal discharge
of someone with bacterial vaginosis, Gram
staining will reveal gram-negative coccobacilli
that are adhered to epithelial cells. Because
bacterial vaginosis does not cause an
inflammatory response, trichomonas or
cervicitis should be suspected if WBCs are
present in large numbers.

Reference(s):

Almassi, E. (2017). What is Bacterial

Vaginosis and How Do Women Get It?
Retrieved from https://nwhn.org/recently-
went-doctor-told-bv-bacterial-vaginosis-
women-get/

Cook, R., Pond, D., Reid, G., Schmitt, C.,

& Sobel, J. (1989). Clue cells in bacterial
vaginosis: immunofluorescent identification of
the adherent gram-negative bacteria as
Gardnerella vaginalis.
DOI: 10.1093/infdis/160.3.490

Money, D. (2005). The laboratory diagnosis of

bacterial vaginosis. doi: 10.1155/2005/230319

Kelly, K. (n.d.). Tests on Vaginal Discharge.

Retrieved from
https://www.ncbi.nlm.nih.gov/books/NBK288/
A woman had her histopathologic Breast cancer tissue removed during a
biopsy is stained using a technique known as
immunohistochemistry, or IHC. IHC is used to
determine whether cancer cells have HER2
receptors and/or hormone receptors on their
surfaces.
IHC is the most commonly used test to
determine whether cancer cells in a tumor have
an excess of the HER2 receptor protein on their
surface. When there are too many HER2
receptors, the cells receive too many signals
instructing them to expand and divide. The IHC
test, which provides a score ranging from 0 to
3+, measures the level of HER2 receptor
protein on the surface of cells in a breast
cancer tissue sample. If the score is 0 to 1+, it
is considered HER2 negative. If the score is 2
or higher, it is considered borderline. A score of
3 or higher indicates HER2 positivity. Many
HER2-negative breast cancers express some
HER2 proteins on the cell surface.
examination in breast tissue after
mastectomy came to the laboratory to
request pull out of her slides and blocks
because her oncologist wants to do a second
opinion on her case. A letter was presented
by the patient duly signed by her physician.
Immunohistostaining was requested and
needs to be sent to PGH. A deposit fee was
suggested and the patient prompted to pay
for the due to proceed with her test. What do
you think is/are the immunohistostaining
technique/s that is/are requested by her
physician? For what purpose do they fit in to
ensure the patient’s good prognosis?
As a medical technologist, what do you
expect the results will be in terms of the
following:
a. Hematoxylin and eosin (H&E) stained
tissue section shows blue blob
patterns
b. Tissue bounces out of paraffin block
during microtomy or tissue does not
adhere to block or slides
c. Tissue does not adhere to slide or
falls off easily
What do you think causes these types of
problem?
Reference(s):
Breastcancer.org. (n.d.). IHC
(ImmunoHistoChemistry) Tests. Retrieved from
https://www.breastcancer.org/screening
testing/ihc-immunohistochemistry-tests
a) If the tissue was properly fixed, the sample
was dehydrated and infiltrated with paraffin
incorrectly. Changing the reagent and
reprocessing the tissue on proper
processing protocol might be needed.
b) Water left in the tissue caused poor
dehydration and paraffin infiltration.
Changing the reagent and reprocessing the
tissue on proper processing protocol might
be needed.
c) Tissue is under-processed when tissue
slides are placed in the oven prior to
deparaffinization in xylene, or when
reagents are saturated with water or
contaminated with the preceding reagent.
Replace reagents and paraffin, and
reprocess tissue according to the proper
processing protocol.
Reference(s):
LabCE. (n.d.). Troubleshooting Processing
Problems. Retrieved from
https://www.labce.com/spg572653_troublesho
25-year-old female came to the hospital due
to painless vaginal bleeding which she said
was heavier than her usual menstrual period.
LMP was 15 weeks ago. A home pregnancy
test kit showed positive. Increase in blood
pressure even without distress was noted.
Elevated β-hCG of 200,000 IU/mL was
recorded. Ultrasonographic imaging showed
intrauterine mass in snowstorm pattern and
an absent fetus is seen. D&C was done to
the patient and chorionic villi was submitted
to the laboratory for examination. Given in
the figure below, describe the gross tissue (1)
immersed in water, and (2) actual tissue
submitted. What is the closest clinical
impression of the patient based on the case
given?
The most common symptom of a molar
pregnancy is vaginal bleeding. A uterine size
greater than expected for gestational age is the
most common physical exam finding of a molar
pregnancy.3 Quantitative beta-hCG levels
greater than 100,000 mlU/mL should raise
suspicion for a molar pregnancy. Molar
pregnancy with normal beta-hCG levels, on the
other hand, is possible.
Ultrasound is the most commonly used
imaging modality for detecting molar
pregnancy. A'snowstorm pattern' has been
described in the past as the result of a complex
vesicular intrauterine mass containing many
'grape-like' cysts. Due to ovarian stimulation
caused by abnormally elevated beta-hCG
levels, ultrasound examination of the adnexa
can also reveal theca lutein cysts.
This is a rare gestational trophoblastic
disease that manifests as a mass of swollen
chorionic villi growing in grape-like clusters. It is
caused by an abnormal egg that has lost its
DNA (deoxyribonucleic acid) and is fertilized by
one or two sperm, resulting in 100% genetic
material from the father.
Reference(s):
Emerg, W. (2013). Ultrasound Detection of a
Molar Pregnancy in the Emergency Department.
doi: 10.5811/westjem.2012.7.12994
Calvo, J. (n.d.). Hydatidiform mole. Retrieved
from
https://www.sciencephoto.com/media/1192182/
view/hydatidiform-mole
Barbieri, R. (n.d.). Gestational Trophoblastic
Disease: Molar Pregnancy. Retrieved from
https://obgynkey.com/gestational-trophoblastic-
disease-molar-pregnancy/
Figure 1.

Figure 2.