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Immunology-Nicole > Immunodiagnostic Questions > Flashcards

Immunodiagnostic Questions Flashcards

1
Q

What 4 substances are considered anti-coagulants?

A
  1. Citrate
  2. Oxalate
  3. EDTA
  4. Heparin
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2
Q

what is the role of an anti-coagulant that has been added to a tube of blood?

A

They all bind calcium

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3
Q

what anti coagulant does not bind calcium?

A

Heparin

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4
Q

why does heparin not bind calcium?

A

Instead it increases the activity of antithrombin

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5
Q

What is plasma in a blood sample?

A

fluid component of uncoagulated blood

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6
Q

what is serum in a blood sample?

A

fluid component of coagulated blood (aka contains clotting factors)

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7
Q

So why is fibrinogen present in plasma and not serum?

A

because proteins that are part of the coagulation system are still present in plasma (aka fibrinogen) and in serum these proteins have been consumed by the formation of the clot .

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8
Q

Simple pneumonic for remembering serum and plasma

A

Serum has less letters than plasma so it has less stuff.

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9
Q

what is the purpose of an indirect immunofluorescence test?

A

detect the presence of a soluble autoantibody

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10
Q

In an indirect immunofluorescence test a small slide contains small slice of frozen tissue of which tissues?

A
  1. Human Thyroid tissue
  2. Murine Kidney
  3. Stomach
  4. Liver
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11
Q

what are you trying to determine with the indirect test and these associated tissues?

A

with patients diluted serum added to the slide the serum will contain autoantibodies if that patient has an autoimmune disorder

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12
Q

If a patient does have autoantibodies reactive with one of the tissues the antigen will bind. However, the autoantibodies can not be visualized. So how do you visualize these auto antibodies?

A

via a tagged fluorescent antibody, anti-human IgG is added (considered a secondary antibody)

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13
Q

what are the 5 disorders for which anti-nuclear antibodies (ANA) may be detected?

A
  1. Systemic Lupus Erythematosus
  2. Rheumatoid Arthritis
  3. Juvenile Chronic Arthritis
  4. Drug Induced lupus ( Hashimoto’s thyroiditis)
  5. Graves Disease

GRADS

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14
Q

ANA antibodies is a screen test not a confirmatory test, why?

A

screens for a number of disorders

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15
Q

what is a direct fluorescence test?

A

adding fluorescent antibodies that will bind to autoimmune antibodies or immune complexes that are already bound to tissue.

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16
Q

why do a direct over an indirect fluorescence test?

A

testing for one specific disorder via a specific antibody

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17
Q

what is the pattern on a renal biopsy for Goodpasture Syndrome?

A

Linear! due to binding to anti-glomerular basement membrane antibodies

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18
Q

what is the pattern on a renal biopsy for Systemic Lupus Erythematosus?

A

Irregular or LUMPY BUMPY (buzz word)

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19
Q

What is an ELISA test used for?

A

to detect antigens or antibodies present in a patients serum.

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20
Q

which ELISA test us used to detect antibody in a patients serum?

A

Indirect ELISA

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21
Q

what is the basic schematic for an indirect ELISA test?

A

Antigen –> Primary antibody —> Secondary Antibody –> Enzyme

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22
Q

Which ELISA test is used for detecting antigen in a patients serum?

A

Sandwich ELISA

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23
Q

what is the basic schematic for a sandwich ELISA?

A

Antibody —> antigen —> antibody —> enzyme

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24
Q

what is the tagged antibody for ELISA test?

A

anti-Ig antibody

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25
Q

what is the underlying principle of electrophoresis?

A

migration of molecules under the influence of an electric field

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26
Q

do molecules migrate in the direct of the electrode bearing the same or opposite charge?

A

opposite

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27
Q

Which molecules migrate faster on an electrophoresis gel?

A

smaller particles

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28
Q

Name the order of migration in terms of fastest to slowest of IgA, IgG, and IgM

A

IgG, IgM, IgA

Go Move (your) Ass

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29
Q

what is a paraprotein?

A

an abnormal immunoglobin (Ig) molecule or fragment of Ig produced by a malignant clone of plasma cells

30
Q

in what two biological samples may paraproteins be detected in a patient with multiple myeloma

A

serum or concentrated urine

31
Q

what is the distinguishing factor of multiple myeloma regarding specificity?

A

all plasma cells secrete antibody of the same specificity (all bind the same epitope) because it is a monoclonal gammopathy

32
Q

what is the distinguishing factor of multiple myeloma regarding isotype?

A

all plasma cells secrete antibody of the same isotope as well (defined by the Fc region of the antibody)

33
Q

Why is immunofixation considered a confirmatory test for multiple myeloma?

A

identifies monoclonal antibodies or components of the antibodies in the serum or concentrated urine.

34
Q

what two cancers result in high concentrations of abnormal monoclonal antibodies or paraproteins?

A

multiple myeloma

waldenstrom macroglobulinemia

35
Q

what is the result of antigen-antibody interaction?

A

indicates the different types of chains—> 1.heavy chains gamma, alpha, mu, or 2. light chains kappa or lambda

36
Q

what does a western blot detect?

A

used to detect or identify specific proteins in homogenized tissues

37
Q

why is ELISA a screening test, but the western blot a confirmatory test?

A

western blot —> detects specific antibodies to several HIV proteins in a patients serum
ELISA –> detects antibodies to one particular proteins (gp120) in a patients serum sample.

38
Q

what do agglutination techniques detect?

A

detecting antibodies or antigens in the patients serum sample

39
Q

what are the three categories which agglutination is categorized?

A

direct, passive and reverse passive

40
Q

what is the difference between direct agglutination and passive agglutination?

A

both detect antibodies in the patients serum. however with passive agglutination particles do not naturally express the antigens on their cell surface.

41
Q

When testing a patient for Hashimoto’s Thyroiditis, what antigen is bound to latex particles? What is the antibody in Hashimoto’s Thyroiditis?

A

Latex particles coated with thyroglobulin are used to detect anti-thyroglobulin antibodies

42
Q

When testing a patient for Rheumatoid arthritis, what antigen is attached to the latex particles? what is the autoantibody in this autoimmune disorder?

A

particles are coated with IgG (bound via Fab to expose Fc region) are used to detect rheumatoid factor, an autoantibody in serum of patients with rheumatoid arthritis.

43
Q

What is the difference between passive agglutination and reverse passive agglutination?

A

Passive –> antibodies in patients serum

Reverse passive agglutination –> detects antigens

44
Q

Why is passive Hemagglutination assay termed “hemagglutination” assay?

A

particle is a tanned red blood cell to which an antigen has been passively bound

45
Q

what is rheumatoid factor?

A

patient have autoantibodies that are specific for the Fc region of IgG in the course of their disease. These autoantibodies are called rheumatoid factor

46
Q

what is the isotype of rheumatoid factor?

A

IgM isotype

47
Q

Name 4 conditions that may result in a false positive test for rheumatoid arthritis?

A

tuberculosis , syphilis, systemic lupus erythematosus and viral hepatitis

48
Q

what does titer refer?

A

The titer is the reciprocal of the smallest dilution producing a positive result

49
Q

If a sample is positive at a dilution of 1/160, how do you express the titer?

A

Titer is 160

50
Q

what antigen is used in the agglutination inhibition test for pregnancy?

A

BhCG (latex particles coated with BhCG)

51
Q

Regarding the reaction step why is the reaction not visualized even though an antigen-antibody complex forms?

A

because the antigen-antibody complexes above are soluble, they do not agglutinate. Therefore, the reaction is not detected

52
Q

Explain why the presence of agglutination in the detection step indicates that the patient is not pregnant?

A

Agglutination occurs when unbound antibodies are still present in the assay (from the rxn step) because there was no BhCG in the urine. Therefore antibodies are available to bind to BhCG coated latex particles

53
Q

What does a direct Coombs test detect?

A

used to demonstrate in vivo coating of red blood cells with IgG antibodies

54
Q

what does an indirect Coombs test detect?

A

demonstrates in vitro reactions between red blood cells and antibodies in a patient’s serum sample

55
Q

what are the 4 pathological conditions for which a physician would request a direct coombs test?

A
  1. Autoimmune haemolytic anemia (premature destruction of a patients red cells)
  2. Erythroblastosis foetalis: newborn red cells express maternal anti-Rh antibodies
  3. Drug induced haemolytic anemia (methyldopa–> HTN med)
  4. A proportion of patients with Chronic Lymphocytic Leukemia (CLL)
56
Q

Explain why an anti-human globulin (generated by another species) is used in the indirect coombs test

A

The antihuman globulin (IgG) plus the antibody-coated cells produce agglutination of RBCs

57
Q

Precipitation–> Explain the conditions required to generate a precipitate?

A

Visible precipitation relies on the relevant concentration of antigen and antibody.

58
Q

Precipitation –> Explain the conditions that lead to a false negative when testing for antibodies in the patient

A

a false negative results occurs if antibody is in excess so it is critical that the patient sample is diluted appropriately. No visible precipitation will occur if antigen is in excess

59
Q

Precipitation–> Explain the conditions that lead to a false negative when testing for antigen

A

A false negative will result if antigen is in excess so it is critical that the patient sample is diluted appropriately. No visible precipitation will occur if antibody is in excess

60
Q

VDRL test–> what is the antigen on the slide to which the patient’s serum is added?

A

extract of the ox heart (diphosphatidyl glycerol/cardiolipin)?

61
Q

VDRL test—> explain the conditions leading to a false negative result

A

When antibodies are in excess. so it is critical to dilute the patient’s serum to avoid false negative results

62
Q

VDRL test –> Name one autoimmune disorder that may result in a false positive result

A

Systemic lupus erythematosus

63
Q

Flow Cytometry–> what is the rationale for performing flow cytometry?

A

useful tool for analyzing molecules on cells that are in suspension

the plots provide information regarding expression of cell surface proteins

64
Q

Flow Cytometry –> Explain the role that fluorescent probes have in flow cytometry

A

antibodies to which a fluorescent probe has been attached are incubated with the cells.

The scattered light and emitted fluorescent probe light are converted to signals that can be stored in the computer for analysis

65
Q

Flow cytometry –> name one cell surface protein to identify (a) all B cells, (b) all T cells, (c) subpopulation of T cells (helper versus cytotoxic)

A

(a) All b cells –> anti CD19 antibodies
(b) All T cells –> anti CD3 antibody
(c) Subpopulation of T cells–> anti CD4 antibody

66
Q

Flow cytometry –> the CD4+ T cell count contributes to the diagnosis of AIDS. What is the normal CD4+ T cell count?

A

500-1000 cells/mm3

67
Q

Flow cytometry –> Interpret flow cytometry plots

A

two parameter plot divided into 4 sections to distinguish populations

(i) lower left quadrant –> negative for both parameters
(ii) upper left quadrant –> positive for y axis parameter but negative for x axis parameter
(iii) lower right quadrant –> positive for x axis parameter but negative for y axis parameter
(iv) upper right quadrant –> positive for both parameters or double positive

68
Q

CH50 test –> why is the Hemolytic titration CH50 assay a screening test instead of a confirmatory test?

A

CH50 test can give a quantitive value for the functional activity of total complement. it does not identify which component(s) are deficient.

69
Q

Name two tests that could be used as a confirmatory tests if the CH50 shows a deficiency in overall complement

A

latex agglutination assay

ELISA

70
Q

Interpreting the results of a CH50 test: How many CH50 units are required for a normal functional activity of Total Classical Complement?

A

101-300

Immunology-Nicole (20 decks)

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