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Hematology > Hematology Exam 3 > Flashcards

Hematology Exam 3 Flashcards

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1
Q

What is the total VOLUME of one side of the hemacytometer? What is the total AREA of one side of the hemacytometer?

A

0.9 mm^3 (volume)
9 mm^2 (area)

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2
Q

How thick is the coverslip placed on top of the hemacytometer?

A

0.1 mm

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3
Q

What is the area of the small “R” squares in a hemacytometer?

A

0.04 mm^2

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4
Q

What is the area of the large “W” squares in a hemacytometer?

A

1 mm^2

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5
Q

What is the calculation for the hemacytometer?

A

Total count (cells/uL) = cells counted x dilution factor / squares counted x area (mm^2) x 0.1

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6
Q

If an average of 96 cells were counted in 4 large squares, what is the total cell count?

A

(20)(96)/(4)(1)(0.1) = 4800 cells/uL

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7
Q

What is the dilution used for manual WBC counts?

A

1:20 dilution

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8
Q

When would you perform a correct WBC count and why? FORMULA FOR THIS WILL BE GIVEN ON EXAM

A

If there is a presence of 5 or more NRBCs a corrected WBC count is needed since NRBCs are not lysed by the diluting fluid, they can falsely increase the WBC count.

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9
Q

The corrected WBC count should always be ______ than the first WBC count.

A

Lower

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10
Q

Which area of a hemacytometer is counted for manual PLT count? What dilution is used?

A

1:100 dilution is used; count the 25 small squares inside of the center large square

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11
Q

Describe the cyanmethemoglobin for determining HGB

A

Blood is diluted in Drabkin’s solution.
Fe2+ Hemoglobin is oxidized to Fe3+ methemoglobin by K ferricyanide found in the solution.
Fe3+ methemoglobin is then converted to Fe3+ cyanmethemoglobin by potassium cyanide found in the solution.
Absorbance of cyanmethemoglobin at 540nm is directly proportional to the hemoglobin concentration.

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12
Q

What is Drabkin’s solution?

A

Used in cyanmethemoglobin method for hemoglobin determination - consists of potassium ferricyanide and potassium cyanide

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13
Q

Hematocrit

A

Volume of pRBCs that occupies a volume of whole blood AKA PCV packed cell volume

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14
Q

Describe the rule of three in hematology. What does it mean if results do not follow this rule?

A

The hematocrit should be 3x the value of hemoglobin (+/- 3). If results do not follow this rule, the patient has abnormal RBCs (hypochromic or microcytic) or there was an error in testing.

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15
Q

Normal range for MCV.
What is the calculation for MCV?

A

Normal: 80-100 fL (<80 = microcytic, >100 = macrocytic)
Calculation: HCT% x 10/RBC count

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16
Q

Normal range for MCH.
What is the calculation for MCH?

A

Normal: 26-32 pg
Calculation: HGB (g/dL) x 10/RBC count

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17
Q

Normal range for MCHC.
What is the calculation for MCHC?

A

Normal: 32-36 g/dL (<32 = hypochromic, >36 = normochromic, spherocytes)
Calculation: HGB (g/dL) x 100/HCT

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18
Q

What is a reticulocyte?

A

Last immature RBC stage

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19
Q

What is the reticulocyte count used for?

A

To assess the erythropoietic activity of the bone marrow

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20
Q

What stain is used for a reticulocyte count

A

New methylene blue

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21
Q

How to identify a reticulocyte during a retic count

A

Two or more particles of blue-stained material (RNA) is observed

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22
Q

How is a manual retic count performed?

A

Count 1000 RBCs under oil and count retics you see. Retic count = Retics/1000 cells counted = Retic %

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23
Q

What do increased reticulocyte counts mean?

A

Could indicate anemia; the bone marrow is trying to release RBCs prematurely to make up for hypoxia

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24
Q

How to use the Miller Disc? What is the purpose?

A

The purpose is to reduce the labor-intensive process of counting 1000 RBCs for reticulocyte counts.
RBCs are counted in the smaller square (B) and reticulocytes are counted in the larger square (A) and should count at least 112 cells in the small square

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25
What is the absolute reticulocyte count? (ARC)
The actual number of reticulocytes in 1L of blood; reported in a number instead of %
26
What is the purpose of the reticulocyte production index? (RPI)
Shift reticulocytes were released from the BM to compensate for anemia and will falsely increase the retic count; so RPI is used to calculate a correction factor
27
What is an ESR
The distance in mm the RBCs fall in 1 hour
28
What is the purpose of the ESR
to detect and monitor the course of inflammatory conditions, infections, or malignancies
29
What factors can elevate ESR?
Rouleaux, increased plasma proteins, pregnancy
30
What factors can decrease ESR?
anemia, sickle cell, polycythemia, newborn
31
What is the purpose of POC testing?
Offers ability to provide rapid and accurate results at the patient's bedside
32
List 3 hematology POC tests
Hematocrit, Hemoglobin, Cell and PLT Counts
33
What two basic principles of operation do most automated analyzers rely on?
Electronic impedance (resistance) and Optical scatter
34
What is the impedance principle?
Based on the detection and measurement of changes in electrical resistance produced by cells as they traverse a small aperture
35
Coulter's Principle of Electrical Impedance: What do the number of pulses represent?
The number of cells counted
36
Coulter's Principle of Electrical Impedance: What does the height of the voltage pulse represent?
Volume of the cell
37
Coulter's Principle of Electrical Impedance: where is the data plotted?
Volume distribution histogram
38
Coulter's Principle of Electrical Impedance: What is the X and Y axis on the volume distribution histogram?
X axis: volume of each cell Y axis: number of cells
39
Coulter's Principle of Electrical Impedance: What separates the cell populations?
Volume thresholds
40
Coulter's Principle of Electrical Impedance: What are the three parts WBCs are separated into?
Lymphocytes, mononuclear cells (monocytes), and granulocytes (eos, basos, neutros)
41
BRIEFLY describe radiofrequency and direct current.
Cell volume is proportional to the change in direct current, cell interior density is proportional to change in radiofrequency signal. Allows for a five-part differential: neutrophils, lymphs, monos, eosinos, basos
42
What does forward angle light scatter correlate with? What does side scatter correlate with?
Forward angle: (0 degrees) correlates with cell volume Side scatter: (90 degrees) correlates with cell complexity
43
Which instrument uses MAPSS to perform its WBC differential?
Cell-Dyn
44
What parameters are affected by Cold agglutinins? Why? How do you correct this?
Decreased RBC and Increased MCV and MCHC due to agglutination of RBCs; correct by warming specimen to 37C and rerun
45
What parameters are affected by Lipemic/icteric specimens? Why? How do you correct this?
Increased HGB and MCH due to increased turbidity affecting spectrophotometric reading; correct with plasma replacement
46
What parameters are affected by hemolytic specimens? Why? How do you correct this?
Increased HCT, Decreased RBC due to RBCs lysed and not counted; correct by requesting new specimen
47
What parameters are affected by PLT clumps? Why? How do you correct this?
Decreased PLT, Increased WBC due to large clumps counted as WBCs and not PLTs; correct by redrawing specimen in sodium citrate and multiply result by 1.1
48
What parameters are affected by old specimens? Why? How do you correct this?
Increased MCV, MPV and decreased PLT due to RBCs swelling as specimen ages and PLTs swell and degenerate. Correct this by establishing stability and specimen rejection criteria
49
In what time frame should blood films be made?
Within 4 hours of collection
50
What is the most commonly used method for making peripheral blood films?
Manual wedge technique
51
How would you adjust the angle of the blood smear for a patient with a high HCT?
Lower angle
52
How would you adjust the angle of the blood smear for a patient with a low HCT (anemic)?
Higher angle
53
What is in included in Wright-Giemsa stain?
Polychrome stains (eosin and methylene blue), giemsa, methylene
54
What is the purpose of methylene in the Wright stain?
Fixes cells to the slide
55
What type of stain is eosin? What does it stain?
It is an acidic stain that stains basic cell components such as HGB and eosinophilic granules
56
What type of stain is methylene blue? What does it stain?
It is a basic stain that stains acidic cellular components such as RNA
57
Why might RBCs appear too gray, WBCs appear too dark, or eosinophil granules appear gray on a blood film?
Stain or buffer was too alkaline, inadequate rinsing, prolonged staining, or heparinized blood specimen
58
Why might RBCs appear too pale/red or why might WBCs be barely visible on a blood film?
Stain or buffer was too acidic, underbuffering, overrinsing
59
Describe the optimal assessment area for examining a peripheral blood smear.
RBCs should be uniformly and singly distributed with few touching or overlapping and have a normal biconcave appearance (~200-250 RBCs per 100x field)
60
SI vs Common units
SI = per L Common = per uL, dL, or %
61
Microcytosis
RBCs are smaller than average (low MCV)
62
Macrocytosis
RBCs are larger than average (high MCV)
63
Anisocytosis
RBCs are unequal in size and have a large variability (large RDW)
64
Hypochromia
Larger central pallor (low MCHC)
65
Polychromasia
Appearance of blue-gray cells with red cells indicating premature release of RBC from bone marrow
66
Poikilocytosis
Increase in abnormally shaped RBC
67
RBC morphology parameters
Cell size (microcytosis, macrocytosis) Variability in size (anisocytosis) Cell color (polychromasia) Cell shape (poikilocytosis) Cellular inclusions (pappenheimer bodies, toxic granulation, etc.)
68
WBC parameters reported in CBC
Total WBC count (WBCs x 10^9/L) WBC differential count (relative) WBC differential count (absolute) WBC morphology
69
PLT parameters reported in CBC
PLT count (PLTs x 10^9/L) Morphology MPV (mean platelet volume)
70
RBC parameters reported in CBC
RBC count (RBCs x 10^12/L) HGB (g/dL) HCT (%) MCV MCH MCHC RDW Morphology
71
A hypochromic, microcytic anemia would have abnormal results for which two main RBC parameters?
Low MCV Low MCHC
72
What RBC parameter is anisocytosis associated with?
Large RDW
73
What RBC parameter is macrocytic associated with?
MCV (high)
74
What RBC parameter is hypochromic associated with?
MCHC (low)
75
What are the main reasons flow cytometry testing is performed?
To detect/monitor leukemias, lymphomas, and HIV
76
What are the proper storage requirements for flow cytometry specimens (blood, bone marrow, fluids, tissues)
Blood and BM: Room temp less than 24hrs Fluids and Tissue: Refrigerated 4C less than 24hrs
77
Monoclonal antibodies in flow cytometry
Antibodies made from one type of cell
78
CDs in flow cytometry
Clusters of differentiation - cluster of antibodies recognizing the same antigen
79
BRIEFLY describe how a flow cytometer works
uses hydrodynamic focusing and lasers as light sources to produce both scattered and fluorescent light signals that are read by detectors, signals are converted into electronic signals that are analyzed by a computer
80
What is gating in flow cytometry?
Selection of a population of interest as defined by one or more flow cytometric parameters
81
What cells show the highest density of CD45 in flow cytometry?
Lymphocytes, then monocytes
82
What cells show intermediate CD45 density?
Granulocytes
83
What cells are negative for CD45?
Late erythroid precursors and megakaryocytes
84
What cells are the most complex (most side scatter)?
Granulocytes
85
What cells are the least complex (not much side scatter)
Lymphocytes
86
What cells are biggest (most forward scatter)
Monocytes
87
What CD markers are present on megakaryoblasts?
CD31 and CD36
88
What CD markers are the first of megakaryocytic differentiation?
CD41 and CD61
89
What are blasts characterized by?
Low density expression of CD45
90
What is the CD4:CD8 ratio in healthy individuals?
>1 (more CD4)
91
What is the CD4:CD8 ratio in HIV positive patients?
<1 (losing CD4)
92
How is flow cytometry used to monitor HIV therapy?
CD4 and CD8 antibodies used to measure CD4:CD8 ratio
93
What RBC, granulocyte, and monocyte CD markers are decreased in PNH?
RBC: CD55 and CD59 Granulocyte: CD24 and FLAER Monocyte: CD14 and FLAER
94
List the purines
Adenine and Guanine
95
List the pyrimidines
Thymine, Cytosine, and Uracil
96
In DNA, A pairs with?
T
97
In RNA, A pairs with?
U
98
In DNA, G pairs with?
C
99
Why is isolation of RNA more difficult than DNA?
There is RNases present on the surface of human skin which can lead to contamination of lab surfaces and inhibition of amplification
100
Regular PCR vs RT-PCR
Regular PCR amplifies DNA, RT-PCR amplifies RNA
101
Process/Purpose of PCR
Denaturation of DNA, DNA polymerase binds to DNA and reads template and amplicon is reproduced millions of times to detect viruses/bacteria genomes
102
What is gel electrophoresis
Separation of DNA fragments by their MASS only
103
Agarose gel vs polyacrylamide gel
Agarose gel has larger pores and allows for larger fragments to pass Polyacrylamide gel has smaller pores and is more effective for smaller fragments
104
Anode vs cathode
Anode is the positively charged electrode; Cathode is the negatively charged electrode
105
How do fragments move in electrophoresis and why
Since DNA is negatively charged, it moves towards the positively charged anode
106
What fragments move further in gel electrophoresis?
Smaller fragments will move faster and at a further distance
107
How does capillary gel electrophoresis differ from regular gel electrophoresis?
Capillary gel electrophoresis is automated and separation is rapid and has greater resolution with more accuracy
108
In one sentence, describe the theory behind RFLP (Restriction Fragment Length Polymorphism)
If there is a mutation present, restriction endonucleases will not cleave the same sites, and will produce restriction fragments of different lengths than normal.
109
Describe real-time PCR Ct/Cp
Ct is cycle threshold and this is the PCR cycle at which amplification crosses the threshold; it is inversely related to the amount of target; the more starting DNA that is present, the lower the number of PCR cycles required to amplify.
110
Describe qualitative vs quantitative real-time PCR
Qualitative PCR is looking for the presence or absence of a mutation while quantitative PCR is looking for the amount of copies of the mutation present
111
What types of body fluids can be tested in the hematology lab?
- CSF - Serous/body cavity fluid (pleural, pericardial, peritoneal fluids) - Synovial (joint) - Bronchoalveolar lavage (BAL)
112
Describe ependymal/choroid plexus cells. Which body fluid can they be found in?
They can be found in CSF because they line the CNS, and they are clumps of basophillic cells with round nuclei
113
Describe cartilage cells. Which body fluid can they be found in?
These are seen in CSF if vertebral body is accidentally punctured - they are a deep wine-red color.
114
Describe siderophages. Which body fluid can they be found in?
These can be found in CSF and they are macrophages that have ingested RBCs and as a result of the breakdown of RBCs contain hemosiderin and sometimes bilirubin. Bilirubin = yellow crystals Hemosiderin = large dark blue/black granules in cytoplasm
115
Describe mesothelial cells. Which body fluid can they be found in?
They can be found in serous fluids (like pleural fluid). They have a fried egg appearance and may be multinucleated.
116
Describe LE cells. Which body fluid can they be found in?
Lupus erythematosus cells may be seen in serous fluids of patients with lupus (SLE). They are intact neutrophils that have engulfed nuclear material, which displaces the normal nucleus.
117
Describe signet ring cells. Which body fluid can they be found in?
Signet ring cells are macrophages that have ingested lipids and can be found in serous fluids. They are a clear-like cell with a purple border and the nucleus on one far edge of the cell.
118
Describe ciliated epithelial cells. Which body fluid can they be found in?
These are normally found in BAL specimens and should be reported because they indicate that the sample was taken from the URT instead of deeper in the lung. They are columnar cells with the nucleus at one end of the cell and cilia at the opposite end of the nucleus.
119
What are the most common cells seen in a BAL?
Neutrophils, monohistiocytes (macrophages), lymphocytes

Hematology (10 decks)

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