Diagnostic Techniques

Question 1

flow cytometry

Answer 1

• usually performed on BM, peripheral blood, or lymphoid tissues
• measures cellular characteristics based on their ability to scatter light (optical scatter)

Question 2

forward scatter

Answer 2

cell volume/size

Question 3

side scatter

Answer 3

cellular complexity

- nucleus, is the cell granulated?

Question 4

immunophenotyping

Answer 4

phenotyping cell surface markers through Abs interacting w antigens (fluorescently-labeled monoclonal Abs)

Question 5

useful in diagnosing hematological malignancies

Answer 5

cell markers

- neoplastic cells will stop at a certain point in maturation and will display aberrant antigenic patterns

Question 6

CD

Answer 6

cluster of differentiation

- cell markers specific for those types of cells like CD4, CD8, etc.

Question 7

CD8

Answer 7

suppressor T cell

Question 8

CD45

Answer 8

leukocyte common antigen

Question 9

CD34

Answer 9

immature hematopoietic cell

Question 10

CD19, CD20

Answer 10

precursor and mature B cells

Question 11

molecule diagnostics

Answer 11

• used to detect specific sequences of DNA/RNA in nucleated blood or BM cells, tissues, or body fluids
• PCR is used to amplify DNA, followed by gel or cap electrophoresis for the detection of amplified DNA
• DNA/RNA in a sample can also be measured quantitatively

Question 12

Reagents needed in a PCR

Answer 12

• primers
• DNA polymerase (Taq)
• nucleotides (dATP, dCTP, dGTP, dTTP)
• Mg buffer
• thermocycler

Question 13

enzyme-based method for amplifying a specific DNA target sequence

Answer 13

Polymerase Chain Reaction

Question 14

Reverse Transcriptase PCR (RT-PCR)

Answer 14

• detects RNA expression
• mRNA used as the starting material
• reverse transcriptase produces complementary DNA (cDNA) from mRNA
• cDNA amplified using specific primers
• method can be paired with fluorescent probes for quantitatively measurement of DNA amplification

Question 15

study of chromosome structure and inheritance

Answer 15

cytogenetics

Question 16

chromosomes arranged and displayed in order of numbered pairs

Answer 16

Karyogram

- chrom 1 longest and shorter and shorter until chrom 22.. then chrom 23 = sex chromosomes

Question 17

human somatic cells have __ chromosomes

Answer 17

46 (diploid or 2n)

**gametes have 23 chromosomes (haploid or n)

Question 18

genes arranged on the chromosomes in humans

Answer 18

25 000

Question 19

3 shapes of chromosomes

Answer 19

• metacentric
• submetacentric
• acrocentric

Question 20

4 stages in the cell cycle

Answer 20

G1 = growth period
S = DNA synthesis
G2 = post DNA synthesis
M = mitosis
want to stop cell division here bc chromatin is thickest and best visualized ; specifically at metaphase (can chemically arrest cells here)

Question 21

G banding

Answer 21

• routinely performed using Giemsa stain

- stains A and T rich areas and do not stain C and G

Question 22

R banding

Answer 22

highlights telomeres

Question 23

Q banding

Answer 23

rapid identification of the Y chromosome

Question 24

C banding

Answer 24

stains the centromeres and condensed heterochromatin

Question 25

Fluorescence In Situ Hybridization (FISH)

Answer 25

- couples molecular method with karyotypes to increase detection of mutations - target DNA is denatured into ssDNA - fluorescent-labeled DNA or RNA probe is used > hybridize to chromatin cells on a glass slide > red and green coloured probes to correspond to different targets on a chromosome - allows for the detection of fusion vs. break-apart signals

Question 26

Acute promyelocytic leukemia

Answer 26

- abnormal proliferation of blasts; very very granulated like promyelocytes - t(15;17)(q24;q21)

Question 27

Acute myeloid leukemia with eosinophilia

Answer 27

inv(16)(p13.2q22)

Question 28

Cytochemistry

Answer 28

- older method - staining of unique cellular components to aid in cell identification - can differentiate cells based on lineage and maturation - cytochemical stain examples: > Myeloperoxidase, esterases, Periodic Acid-Schiff, acid phosphatase

Question 29

Periodic Acid Schiff

Answer 29

- reacts with glycogen, mucoproteins, and high molecular weight carbs - positive reactivity: > leukemic lymphoblasts (block positivity) > leukemic erythroblasts (coarse and granular) > granulocytes (+ increases w maturity) > megakaryocytes (diffuse positivity)

Question 30

Acid Phosphatase

Answer 30

- various isoenzymes of acid phosphatase are found in all hematopoietic cells - lymphocytes in hairy cell leukemia produces an isoenzyme resistant to tartrate > tartrate-resistant acid phosphatase (TRAP stain)

Question 31

Acid Phosphatase

Answer 31

- various enzymes of acid phosphatase are found in all hematopoietic cells - lymphocytes in hairy cell leukemia produces an isoenzyme resistant to tartrate > tartrate-resistant acid phosphatase (TRAP stain)

Question 32

gel electrophoresis

Answer 32

- detection of DNA - DNA fragments migrate through gel - rate of migration based on size and weight - agarose or polyacrylamide gel used - smaller fragments move the farthest - many different methods of measuring DNA fragments - fluorescent dyes used to visualize fragments under UV light - qualitative measurement of DNA

Question 33

The chromatin condenses during _______ to form ___________

Answer 33

mitosis; chromosomes

Question 34

aneuploidy

Answer 34

- numerical chromosomal abnormality | - not a multiple of 23

Question 35

polyploidy

Answer 35

- numerical chromosomal abnormality | - multiple of 23, greater than 46

Question 36

chromosomal structural abnormalities

Answer 36

- rearrangements of genetic material - may be balanced or unbalanced - due to inversions, deletions, translocations, or duplications

Question 37

myeloperoxidase

Answer 37

- usuall found in primary granules of neuts and eos; basos have decreased + with maturity; monocyte variable - not present in lymphs - leukemic myeloblasts are positive (differentiate from lymphoblasts)

Question 38

esterases

Answer 38

- differentiates leukemic cells that are granulocytic vs monocytic - substrates: alpha-naphthyl acetate (NSE) = monos are positive; positivity inhibited after adding NaF - naphthol AS-D chloroacetate (SE) = granulocytes stain positive; monocytes stain weak to negative