Quiz 1 Review ( Immunoassays)

Question 1

What are immunoassays?

Answer 1

Tests that are based on antigen-antibody reactions

Question 2

Immunoassays are used to measure what concentrations of analytes?

Answer 2

< 1 μmol/L

Question 3

The region of the antigen that binds to a complementary antibody is
called?

Answer 3

An epitope

Question 4

The strength of a single antibody-antigen interaction is called?

Answer 4

Affinity

Question 5

The combined strength of all antibody-antigen interactions is called?

Answer 5

Avidity

Question 6

What is cross-reactivity?

Answer 6

The ability of an antibody to react with more than one epitope.
(i.e. Ab reacts with structurally similar antigens)

Question 7

Which is more sensitive labeled or unlabeled immunoassays?

Answer 7

Labeled

Question 8

What types of labels can be used in labeled immunoassays?

Answer 8

Radioactive element
Fluorochrome
Chemiluminescent compound
Enzyme

Question 9

What are the common chemiluminescent labels?

Answer 9

Luminol

Acridinium esters

Question 10

What are the most common enzyme labels?

Answer 10

Alkaline phosphatase (ALP)
Horseradish peroxidase (HPO)

Question 11

What is the principle of competitive immunoassays?

Answer 11

Unlabeled antigen in the patient sample competes with labeled reagent antigen for limited binding sites on a reagent antibody. (Equal competition)

Labeled Ag is inversely proportional to patient analyte concentration.

The more unlabeled (patient) antigen in the sample, the less labeled antigen will be bound.

Question 12

What is the difference between heterogeneous and homogeneous
immunoassays?

Answer 12

Heterogeneous immunoassays require a separation step of antigen bound to
antibody from unbound antigen.

Homogeneous immunoassays do not require a separation step.

Question 13

What are some examples of competitive, heterogeneous immunoassays?

Answer 13

RIA - radioimmunoassay
EIA - enzyme immunoassay
FIA - fluorescence immunoassay
ELISA - enzyme-linked immunosorbent assay

Question 14

What are the steps in an ELISA immunoassay to detect an antibody?

Answer 14

1. Patient sample is added to a microtiter well containing reagent antigen. If
   antibody is present in the sample, it will bind to the reagent antigen on the well.
2. A wash step removes any unbound sample.
3. Enzyme-labeled antibody is added and will attach to the patient antibody if
   present.
4. A wash step removes any unbound labeled antibody.
5. Enzyme substrate is added.

Question 15

In a competitive immunoassay, what relationship exists between the
unlabeled patient antigen and the bound labeled antigen?

Answer 15

The more unlabeled patient antigen in the sample, the less bound labeled antigen.

This will mean less radioactivity, color, fluorescence, chemiluminescence, etc.
Inversely related.

Question 16

What are some examples of competitive, homogeneous immunoassays?

Answer 16

EMIT - enzyme-multiplied immunoassay technique

FPIA - fluorescence polarization immunoassay

Question 17

How do non-competitive immunoassays differ from competitive

immunoassays?

Answer 17

Non-competitive immunoassays use an excess of labeled antibody.

The concentration of patient antigen is directly proportional to the bound,
labeled antibody.

Question 18

What are some examples of non-competitive immunoassays?

Answer 18

IRMA - immunoradiometric assay

IEMA - immunoenzymatic assay

Question 19

What can cause false-positive reactions with sandwich immunoassays?

Answer 19

Heterophile antibodies

HAMA

Question 20

What can cause false-negative results when using sandwich

immunoassays?

Answer 20

The hook effect

Occurs when analyte being measured is in very high concentrations. There is
insufficient binding sites for all of the antigen in the sample.

Question 21

How can the hook effect be corrected?

Answer 21

Dilute the sample and rerun.

Question 22

Which immunoassay has a direct relationship between the
concentration of drug in a patient sample and the activity of free
labeled drug?

Answer 22

EMIT

Question 23

In fluorescence polarization immunoassays (FPIAs), how does the light
emission of bound labeled antigen differ from that of free labeled
antigen?

Answer 23

Bound labeled antigen rotates more slowly and emits more polarized florescence.

Free labeled antigen rotates more quickly and emits less polarized florescence.

No separation step required

Question 24

What are some examples of non-labeled immunoassays?

Answer 24

Nephelometry

Turbidometry

Precipitation methods

Question 25

What is the zone of equivalence in a precipitation test?

Answer 25

The point in an antigen-antibody reaction where the number of antigen site and
antibodies are approximately equal.
Optimum precipitation occurs in the zone of equivalence

Question 26

What are examples of passive precipitation methods in gel?

Answer 26

Double diffusion (Ouchterlony)

Radial immunodiffusion (RID)

Question 27

What is a pattern of identity in the Ouchterlony technique?

Answer 27

Single, continuous precipitin arc forms meaning antigens in the specimen and in the control are identical.

Question 28

What is a pattern of identity in the Ouchterlony technique?

Answer 28

Single, continuous precipitin arc forms meaning antigens in the specimen and in the control are identical.

Question 29

When is immunofixation electrophoresis (IFE) used?

Answer 29

To identify immunoglobulins and Bence Jones protein.

Question 30

What technique measures protein based on light scatter by immune
complexes?

Answer 30

Nephelometry

Question 31

What type of specimen could cause inaccurate results in the above
method?

Answer 31

Lipemic specimens would cause light scattering.