Antigen/ Antibody Interactions

Question 1

What causes precipitation of immune complexes? What point of the precipitin curve has optimal lattice formation?

Answer 1

Lattice formation into an insoluble complex; equivalence zone

Question 2

What is the titer in agglutination?

Answer 2

The reciprocal of the most dilute solution at which cross-linking of a particulate antigen is seen.

Question 3

What is the prozone of an agglutination curve?

Answer 3

The part of the precipitation curve in which too much antibody inhibits cross-linking of particles

Question 4

What is the difference in precipitation and agglutination?

Answer 4

Precipitation occurs typically with smaller, initially soluble antigens, while agglutination occurs with larger antigens

Question 5

How is slide agglutination used for diagnosis?

Answer 5

A slide can be coated with an antibody to or antigen of a particular pathogen and then patient sample is added. If agglutination occurs then the patient is positive for the pathogen

Question 6

What is hemagglutination? What is its utility?

Answer 6

Agglutination of RBCs, which can be used to type blood based on the ABO/ Rh blood type

Question 7

What is an isohemagglutinin?

Answer 7

An antibody (mostly IgM, some IgG) that agglutinates red blood cells from a member of the same species

Question 8

How does an hemagglutination inhibition assay work? What is it used for?

Answer 8

Method used to detect infection with influenza virus. The virus has hemagglutinin protein which binds to RBCs- virus can agglutinate RBCs. Patient serum is incubate with the virus- if the patient has made antibody to virus it will bind to the virus and inhibit the binding of the virus to the RBCs.

Question 9

How is the titer of a hemagglutination inhibition assay dtermined?

Answer 9

The titer is the reciprocal of the highest dilution in which inhibition (no agg.) is seen

Question 10

How can hemagglutination inhibition assays be used to test vaccine efficacy?

Answer 10

Antibodies made against previous season’s virus are tested against the current year’s virus. A new vaccine must be developed if HI titers differ by more than 2

Question 11

What is the difference in between a direct and indirect immunoassay?

Answer 11

Direct assays involve a detection antibody binding directly to antigen. Indirect assays involve a detection antibody binding to another antibody bound to antigen

Question 12

What are the benefits of an indirect immune assay?

Answer 12

An indirect assay amplifies the response when a direct assay is not sufficient to visualize results, it is more cost-effective too

Question 13

What is a Coombs Test?

Answer 13

Measures for the presence of antibodies to rhesus antigen (D antigen)

Question 14

How is a direct Coombs tests performed?

Answer 14

RBCs are taken from the fetus and determine if they agglutinate with rabbit anti-human Ig

Question 15

How is an Indirect Coombs Test performed?

Answer 15

Serum is taken from mother and incubated with Rh+ RBC, add rabbit anti-human Ig- if positive- agglutination

Question 16

How is a diffusion assay performed?

Answer 16

Soluble antigens and/or Ab are added to wells in agar and allowed to diffuse towards each other to form an equivalence zone

Question 17

How is radial immunodiffusion performed? For what is it used?

Answer 17

Ag s placed into an Ab-agar, Ag diffuses radially and precipitates at the equivalence zone such that the ring diamter is proportional to Ag concentration; Used to quantitate serum proteins

Question 18

What is ouchterlony?

Answer 18

A double diffusion technique where Ag and Ab are placed in separate wells in a gel and can be used to identify cross reacting antibodies or antigens that share epitopes

Question 19

How is immunoelectrophoresis performed?

Answer 19

Antigens are loaded into agar, subjected to electrical current, and migrate based on size and charge. Antiserum is added to the trough underneath and precipitation curves indicate interaction

Question 20

How is a radioimmunoassay performed?

Answer 20

Radio- or fluoro- labeled Ag are used to measure the amount of antigen in a sample

Question 21

What is ELISA? Is direct or indirect more common?

Answer 21

Enzyme-linked immunosorbent assay

Question 22

What is a sandwich ELISA?

Answer 22

Antibody is coated at the bottom of well, antigen is added, and another antibody is added that recognizes the antigen, followed by a labeled anti-Ig antibody

Question 23

Which method of ELISA is most sensitive?

Answer 23

Sandwich ELISA

Question 24

How does flow cytometry work?

Answer 24

Cells are stained with fluorochrome label antibodies and drip one at a time through a laser that produces differing light refraction patterns depending on the cell type

Question 25

For what do complement fixation assays test?

Answer 25

Tests for specific antigens or antibodies in a patient’s sample

Question 26

How does the complement fixation assay work?

Answer 26

Patient’s serum is mixed with an antigen and a standardized amount of commercial complement. If a patient has proper antibody then it will form immune complexes and fix the complement. Serum is then added to sheep RBCs– if lysis occurs that means there was still complement in the serum, meaning there was no antibody. If there is no lysis that means all of the complement was fixed by immune complexes