3. Peripheral Blood Smear Flashcards
The patient demonstrates platelet clumping in his blood (lavender top). When CBC was run, the results showed that his PLT count is low (90,000/mm3) and his WBC count is high (15,000/mm3). The phlebotomist recollected his blood using 3.2% Sodium citrate tube and another CBC was run. New results are 115,000/mm3 and 11,000/mm3, respectively. What set of values should finally appear on this patient’s chart?
Plt Ct: 126,500/mm3
WBC ct: 12,100/mm3
Sources of specimens for Peripheral blood smear
EDTA blood
Anticoagulant-free blood
Blood smears using EDTA spx should be made within ___ after collection
2 hours
Advantages of EDTA blood smear (3)
Multiple blood smears
Blood smears may be prepared later
Prevents plt clumping
Disadvantages of EDTA blood smear
Platelet satellitosis
EDTA-induced platelet clumping
Platelet satellitosis aka
Platelet satellitism
Platelet rosette
What CBC parameter is affected by platelet satellitosis
PLT Ct: pseudothrombocytopenia
Why is there pseudothrombocytopenia in platelet satellitism
When platelets adhere to WBCs’ surface - they are also counted as WBC
In platelet rosette, platelets usually adhere in what type of WBC
Neutrophils
How to correct platelet satellitosis and edta-induced platelet clumping
recollect using 3.2% sodium citrate; multiply the affected parameters by 1.1
Correction factor when using 3.2% sodium citrate is
1.1
Why is there a need for a correction factor when recollecting a specimen using 3.2% sodium citrate
There is a need to compensate for the dilution by Na citrate; since there is too much AC
In EDTA-induced platelet clumping, platelets form large clumps and are counted as
WBCs
In EDTA-induced platelet clumping, platelets form large clumps and are counted as WBCs, therefore the affected CBC parameters are
PLT count: pseudothrombocytopenia
WBC count: pseudoleukocytosis
T/F
There is a need to correct all the CBC parameters using the correction factor after recollecting using the light blue top with 3.2% Na citrate
False
Anticoagulant-free blood is rarely used in the hematology section because of (2) disadvantages
Platelet clumping
Few films can be made
(2) advantages of using anticoagulant-free blood for PBS
made at the patient’s side
some artifacts may be prevented
(3) Methods of blood film preparation
Two-glass slide method
Coverslip technique
Automated methods
Two-glass slide method is also known as
Manual Wedge Technique
The most frequently used method in the preparation of blood smear
Manual Wedge Technique/Two-glass slide method
How many slides are used in manual wedge method?
2: pusher slide and film slide
Pusher slide is also known as
spreader
The angle between the pusher slide/spreader and the film slide should be
30-45 degrees
If the angle of the spreader is too high, the smear is expected to be
thicker
If the angle of the spreader is too low, it is expected that the smear is
thinner
The distance between the labeled end and the drop of blood should be
1 cm
The size of blood on the blood smear should be
2-3 mm
If the size of blood in the blood smear is too large, the smear is
thicker and longer
If the size of the drop of blood on the smear is too small, the smear is
thinner and shorter
When the speed of the spreader is too fast, the blood smear is
thicker
When the speed of the spreader is too slow, the smear is thinner with
poor WBC distribution
What makes a blood smear thicker?
too large size of the drop of blood
too fast spreader
too high angle of spreader
What makes a blood smear thinner
too small size of the drop of blood
too slow spreader
too low angle of spreader
In Polycythemia vera, the hematocrit is
too high
If the HCT is too high, the angle of the spreader and the film slide should be as low as
25 degrees
If the HCT is too low i.e. anemia, the angle of the slides should be
raised
Scanning method for examining PBS
Longitudinal
Battlement
Longitudinal pattern
Tail to head
Battlement pattern
back-and-forth serpentine
scanning method utilized for manual wedge technique
longitudinal
ideal blood smear length
2/3 to 3/4 the length of the film slide
Problem encountered in wedge method
uneven distribution of wbcs in the slide
when utilizing the wedge method, the feathered edge and side edges contain more
monocytes, eosinophils, neutrophils
When utilizing the wedge method, the center part of the film contains more
small lymphocytes
(2) types of coverslip technique
glass slide-coverslip method
two-coverslip method
glass slide-coverslip method is also known as
beacom’s method
two-coverslip method is also known as
ehrlich’s method
Beacom’s method and Ehrlich’s method are rarely used. They are sometimes used for
bone marrow aspirate smear preparation
There is only one advantage when employing the beacom’s method and ehrlich’s method
excellent wbc distribution
4 machines in automated method of blood smear preparation
Miniprep
Centrifugal (spinner) type
Coulter LH
Sysmex LP-10
This is a semi-automatic, portable instrument that simulates the manual wedge technique of blood smear preparation.
Miniprep
Miniprep simulates what type of staining method
manual wedge technique
This type of machine uses approximately 0.2 mL of well-mixed anticoagulated blood
Centrifugal (spinner) type
Centrifugal (spinner) type makes use of ______ of well-mixed anticoagulated blood
0.2 mL
What type of automated machine for blood smear preparation is helpful for medical technologists working on field
Miniprep
What type of automated machine for blood smear preparation is useful for conditions like CLL
Centrifugal (spinner) type
In centrifugal (spinner) type, there is a monolayer of evenly distributed cells; and fewer ______ in patients with chronic lymphocytic leukemia
smudge cells
These cells are nuclear remnants of lymphocytes
smudge cells
smudge cells are often mistaken as
debris
These machines are considered as both slide makers and slide stainers
Coulter LH
Sysmex SP-10
Sysmex SP-10 and Coulter LH are both
slide makers
slide stainers
Purpose of blood smear staining
for evaluation of cellular morphology
fixative used in blood smear staining
methanol
stain used in blood smear
wright or wright-giemsa
a chemical that maintains the pH within a particular range
buffer
buffer used in blood smear preparation
0.05 M sodium phosphate
alternative buffer if there is no 0.05 M sodium phosphate
aged distilled water placed in a glass water bottle for at least 24 hours, with pH 6.4-6.8
pH of 0.05 M sodium phosphate
6.4
the correct pH for blood smear preparation
6.4-6.8
Defined as any stain which contains methylene blue (and/or its products of oxidation) and a halogenated fluorescein dye (commonly eosin b or eosin y)
romanowsky-type stain
methylene blue is a basic dye and colors the ____ blue or purple
nucleus and some cytoplasmic structures
eosin stains acidophilic cytoplasmic structures an _____ color
orange-red
the most commonly used type of stain in the hematology laboratory
romanowsky-type stain
Wright stain, wright-giemsa, and May-Grunwald are examples of what type of stain
romanowsky
(3) techniques of staining
manual
automated
quick
Manual staining takes _____ mins
1-3 minutes
Automated staining takes _____ to stain a batch of slides
5-10 minutes
4 machines employed for automated staining
Midas III
Hema-tek
Coulter LH
Sysmex SP-10
Quick staining takes _____ minute/s
1 min
Quick staining uses _____ filtered into a staining dish/coplin jar
modified wright or wright-giemsa stain
What type of buffer is used in quick staining
aged distilled water
ideal macroscopic characteristic of a blood smear
color: pink to purple
Color of RBCs
orange to salmon-pink
color of wbc nuclei
purple to blue
color of neutrophil cytoplasm
pink to tan (with violet to blue granules)
color of eosinophil granules
bright orange
the most common cause of wbcs appearing dark and the rbcs & eosinophil granules appearing gray
stain/buffer is too basic
T/F
Inadequate rinsing leads to wbcs appearing too pale.
False
T/F
Heparin is an ideal anticoagulant in the hematology lab since it does not cause any problems during the staining process.
False
Heparinized blood causes the eosinophil granules to appear
gray
The possible reasons for rbcs appearing too pale or red and barely visible wbcs include:
too acidic stain/buffer
underbuffering
over-rinsing
Probable reason for a blood smear appearing bluer than normal
patient has increased blood proteins
Old name of plasma cell myeloma
multiple myeloma
Multiple myeloma is a condition where a patient excretes bence jones proteins in the urine. In this condition, the PBS of the patient is expected to appear
bluer than normal
A grainy blood smear is probably due to
RBC agglutination
Cold hemagglutinin diseases can cause the blood film to appear
grainy
A patient with increased lipid levels will probably have a blood film that has
holes all over the film
Blue specks at the feathered edge of the blood film indicates that the patient has
increased WBC count
increased PLT count
10x objective is used to (6)
assess overall film quality, color, distribution of cells
locate rare abnormal cells
detect snowplow effect
detect fibrin strands
recognize rouleaux formation
detect parasites
This refers to the presence of >4x the number of WBCs per field at the feathered edge compared with the monolayer area.
Snowplow effect
If fibrin strands are present, the specimen should be rejected since it indicates that
the specimen has already clotted
What area of the blood film should be examined microscopically
the are between the thick part and the feathered edge
There is a need to avoid the feathered edge of the smear when examining since the
RBCs appear macrocytic
WBCs appear distorted
There is a need to avoid the thick part of the blood film since the
RBCs appear microcytic
RBCs seem to form rouleaux
Signs that the area being examined in the blood film is the correct one
RBCs have central pallor, near each other, never overlapping
cells are appropriately stained
This magnification is employed to estimate the total WBC count
40x High-Dry/50x OIO
WBC count estimate formula (40x High-Dry)
average # of wbcs (10 fields) x 2000
WBC count estimate formula (50x OIO)
average # of wbcs counted (10 fields) x 3000
Calculate the estimated wbc count if the average number of white blood cells counted is 5 (40x High-Dry)
10,000/uL
Calculate the estimated wbc count if the average number of white blood cells counted is 5 (50x OIO)
15,000/uL
40x High-Dry/50x OIO is used to get the
estimated total wbc count
range of RBCs per 100x OIF in a normal person
200-250 RBCS per 100x OIF
100x OIO is used to
examine the nuclear details of WBCs
tabulate the actual WBC differential count
platelet count estimate
Formula for platelet estimate (normal)
average # of platelets/OIF (10 Fields) x 20,000 = estimated plt ct/uL
Formula for platelet estimate (anemia, erythrocytosis)
average # plts per field x total RBC count
________________________________
200 RBCs per field
The average number of RBCs per OIF in the optimal assessment area
200
Parasites that may appear in the blood film
Malaria
Filaria
Trypanosomes
Species that can cause malaria in humans
P falciparum
P vivax
P ovale
P malariae
P knowlesi
Most pathologic plasmodium spp
Plasmodium falciparum
most prevalent plasmodium spp
plasmodium vivax
people with ______ seem to be resistant to plasmodium falciparum infections
sickle cell trait
sickle cell trait persons are resistant to
plasmodium falciparum infections
Malarial parasites metabolize this protein to make a malaria pigment.
hemoglobin
Malarial parasites metabolize hemoglobin to produce this malarial pigment
hemozoin
How many blood films should be made to check for malarial parasites ASAP after blood collection
2 thick films
2 thin films
T/F
Blood films can also be directly made from capillary puncture
True
For the visualization of malarial parasites, this type of stain is used
Wright-Giemsa stain
This type of blood film is ideal for the initial screening of blood
thick film
Thick blood film is used to view more parasites. It is stained with water-based Wright-Giemsa stain to
lyse the red cells
The water-based Wright-Giemsa stain done in thick blood films for malarial parasite is done without
methanol fixation
Thin blood films are used for
parasite identification
determination of percent of parasitemia
Thin blood films are stained after
methanol fixation
Degree of parasitemia is determined by counting the number of parasitized RBCs (asexual stages) among — RBCs on a thin blood film and converting it to a percentage
500-2000
Before a negative malaria result is released, how many fields of thick and thin films should be examined
at least 300 (100x OIO)
Microscopy can detect _____ per microliter of blood
5-20 parasites
Microscopy can detect 5 to 20 parasites per uL of blood or
0.0001% parasitemia
T/F
A negative result for a single set of thick and thin PBS rules out a diagnosis of malaria
False
This may be confused with a malarial parasite by an inexperienced observer
platelet lying on top of an RBC
How to prepare thick blood film
3 drops of blood close together
stir for 30 seconds using a clean slide to mix
the drop should be 1-2 cm
Dry
Stain with water-based wright-giemsa
Filaria spp
Wuchereria bancrofti
Brugia malayi
Loa loa
Species that can cause elephantiasis
Brugia malayi
Wuchereria bancrofti
Loa loa can cause
calabar swelling
Trypanosomal spp that infect humans
Trypanosoma brucei rhodesiense
Trypanosoma brucei gambiense
Trypanosoma cruzi
African sleeping sickness vector
Tsetse fly
East African Sleeping Sickness
Trypanosoma brucei rhodesiense
Trypanosoma brucei gambiense is also known as
West African Sleeping sickness
Trypanosoma cruzi can cause
Chagas disease
The feces of this triatomine bug can cause chagas disease
Reduviid bug
Blood smear slides can be stored for at least _____ before disposal
7 days
