Ancillary Diagnostics

Question 1

What is flow cytometry? What can we tell from forward scatter? Side scatter? Fluorescent markers?

Answer 1

Classification of cells by measuring their shape and internal content (granules, hemoglobin etc.)

Forward scatter - size of cells

Side scatter - internal complexity of cells

Fluorescent markers - used to mark molecules that are normally expressed in nucleus, cytoplasm, or membrane of cells

Question 2

What kind of samples can you submit for flow cytometry? How do you submit blood for flow cytometry?

Answer 2

Blood: 500 ul EDTA, cold

Effusions

Aspirates

Question 3

What are some sample limitations for flow cytometry? In other words what features of a sample (sample type, cellularity) will make it unsuitable for flow cytometry testing?

Answer 3

Requires living cells in media or saline

Question 4

What are some general uses of flow cytometry in veterinary medicine?

Answer 4

Further classifications of leukemia

Cause of extreme lymphocytosis

Diagnosing neoplasia/prognostic info

Question 5

Be able to correctly associate the cell markers (CD markers) listed in the notes, with the types of cells that should normally express them.

Answer 5

CD 3 - all T cells
CD 5 - all T cells

CD 4 - T - helper cells

CD 8- cytotoxic T cells

CD21 - B cells

CD34 - precursor cells, non-lymphoid and lymphoid cells

CD 45- all leukocytes

Question 6

If given flow cytometery data from a patient sample, be able to correctly interpret
the results.

Answer 6

.

Question 7

What is PARR testing? What information can be gained by this methodology that cannot be gained by flow cytometry?

Answer 7

PCR or Antigen Receptor Rearrangement

Can confirm clonality of B or T cells

Used in cases of lymphoid malignancy only

Question 8

What samples can you submit for PARR analysis?

Answer 8

EDTA Blood (200 ul)
Stained slides
Effusion/cytology

(Does not require living cells)

Question 9

Be able to identify instances in which PARR analysis would be the preferred diagnostic test compared to flow cytometry.

Answer 9

Do not have a sample with living cells

Suspect lymphoid malignancy with heterogenous cell population

Question 10

Be able to identify instances in which flow cytometry would be the preferred diagnostic test compared to PARR analysis.

Answer 10

Preferred over PARR, but needs living cells

Question 11

Be able to recognize differentials for lymphocytosis in the dog and cat other than lymphoid neoplasia.

Answer 11

Dog:
E. Canis (CD 8), HW, spirocera lupi, leishmaniasis, hypoadrenocorticism, thymoma

Cat: Mycoplasa, toxoplasma, FIV, hyperthyroidism, thymoma

Question 12

Have a general understanding of the cell markers and prognosis associated with the chronic leukemias and T-zone lymphoid neoplasia, as compared to
intermediate to large cell lymphoma, and the acute leukemias.

Answer 12

Chronic B cell leukemia: small cells, CD 21

Chronic T cell leukemia: CD 8

T-zone lymphoid neoplasia: LACK CD 45

B cell lymphoma: large cells, CD 21

T cell lymphoma: CD 4

Acute leukemia: CD 34

Question 13

Be able to list or identify some cell types that can be infected with FeLV.

Answer 13

Lymphocytes

Monocytes/macrophages

Question 14

What are 3 possible outcomes of FeLV infection? What test results would you expect (ELISA/IFA and PCR) in
patients with these different outcomes?

Answer 14

Abortive infection: test negative

Regressive infection: will only test positive on PCR

Progressive infection: will test positive on PCR, ELISA, IFA

Question 15

Be able to list or identify 3 possible causes of regenerative anemia in FeLV infected cats.

Answer 15

1. Hemolytic anemia from secondary infection
2. IMHA
3. Anemia of blood loss

Question 16

Be able to list or identify causes of non-regenerative anemia in FeLV infected cats.

Answer 16

1. Pure red cell aplasia
2. Anemia of inflammatory disease
3. Anemia from bone marrow crowding (myelophthisis)

Question 17

Be able to list or identify some hematologic diseases that are associated with FeLV infection

Answer 17

Pancytopenia
Thrombocytopathy
Leukemia
Erythrolukemia
Myelodysplastic syndrome
Myelofibrosis

Question 18

Can an animal with hematologic disease from FeLV test negative for FeLV by antibody testing? If so why? Is this more common with certain FeLV diseases manifestations?

Answer 18

Yes

Animals with regressive FeLV may have severe, non-regenerative anemia, myelosupression, non-lymphoid hematopoietic malignancies

Depends on place of insertion of proviral DNA