antigen antibody reactions

Question 1

What governs antigen-antibody reaction?

Answer 1

The law of mass action

Question 2

What is the equation showing an antibody-antigen reaction?

Answer 2

[Ag] + [Ab] -> (k1 + k2) [AgAb]

Question 3

What is an application of the antigen-antibody reaction?

Answer 3

It is used in vitro to test for antibodies & antigens

Question 4

How is the antigen-antibody reaction visualised in transfusion science?

Answer 4

Haemagglutination & haemolysis of RBCs

Question 5

How many stages are there in haemagglutination?

Answer 5

2

Question 6

Explain the 1st stage of haemagglutination.

Answer 6

~Known as sensitisation, binding, association, or coating
~Antibodies attach to corresponding antigens
~Antibody combining sites (FAB region) - complimentary in shape & size to antigens - react specifically and form reversible bonds
~bonds in antibody binding include - ionic bonds, hydrogen bonds, hydrophobic bonds, van der Waals,

Question 7

Explain the second stage of haemagglutination.

Answer 7

~Agglutination
~Intracellular bridges form - antibodies already bound to antigens on one RBC bind to the antigens of others RBCs
~Aggregating forces - surface tension change pulls RBCs together
~Repulsive forces - RBC surface - net - charge due to sialic acid & neuraminic acid - prevents RBCs from adhering to each other

Question 8

What are Igs composed of?

Answer 8

~2 heavy chains, 2 light chains
~Variable region & constant region
~Heavy chains bound by disulphide bonds
~Y shaped
~2 Antigen combining bond sites on variable region

Question 9

What is the zeta potential?

Answer 9

Net strength of forces that exist between cells suspended in saline

Question 10

For direct agglutination to occur what distance must an antibody span & what causes it?

Answer 10

20nm cause by zeta potential

Question 11

What is the maximum gap between binding sites for IgMs?

Answer 11

30nm - allows direct agglutination

Question 12

What is the maximum gap between binding sites for IgGs?

Answer 12

14nm - can’t cause direct agglutination

Question 13

What factors affect antigen-antibody reactions?

Answer 13

Temperature, pH, ionic strength, antigen & antibody conc.

Question 14

How does temperature affect that antibody-antigen reaction?

Answer 14

React differently at different temperatures - ABO @ RT , Rh @ 37°

Question 15

How does pH affect that antibody-antigen reaction?

Answer 15

Optimum 6.5-6.8
Little effect between 5.5-8.5

Question 16

How does ionic strength affect that antibody-antigen reaction?

Answer 16

Low ionic strength solutions - more mobile & favour ionic binding between Ig & antigen - speeds up haemagglutination - e.g. 0.8-0.95% saline

Question 17

How does excessive antigens affect that antibody-antigen reaction?

Answer 17

Antibodies are dispersed thinly - formation of intracellular bridges sparse and need too much energy - e.g. RBC cell suspension too strong

Question 18

How does excess antibody affect that antibody-antigen reaction?

Answer 18

Antibodies bind to antigens freely - lack of intracellular bridges as antigens are bound with individual antibodies - (e.g. weak cell suspension)

Question 19

What is direct agglutination in the transfusion lab?

Answer 19

Direct agglutination of red cells suspended in saline is only observed with IgM antibodies which are large enough to span the gap between adjacent red cells

RBCs in saline kept apart @ 20nm - IgM antibodies - large enough to span the gaps between RBCs - ABO grouping - conc. of antigen & antibody optimised - 1/2 plasma/serum : 1 RBC suspension in saline

Question 20

What is indirect agglutination?

Answer 20

When the IgG antibody is too small to cause direct agglutination in vitro
IgG antibody isn’t large enough to form intracellular bridges between red cells.
Therefore another molecule is added to help bridge this gap called anti-human globulin or Coombs reagent.

Anti Human Globulin test - IgG antibodies - incomplete antibody as does not form intercellular cellular bridges - AHG/Coomb’s reagent added to help bridge gap - Coombs test

Question 21

What must be optimised for direct agglutination to occur?

Answer 21

Concentration of antigen and antibody

Mix 1-2 volumes of plasma/serum and a volume of 3-5% suspension of red cells in saline