antigen – antibody reaction

antigen – antibody reaction: 

reaction of antigens and antibodies are highly specific. an antigen will react only with antibodies elicited by itself or by a closely related antigen. 

because of the great specificity, reactions between antigens and antibodies are suitable for identifying one by using the other. this is the basis of serologic reactions. 

antigens: 

are foreign substances, usually high-molecular weight proteins or carbohydrates, that elicit the production of other proteins, called antibodies, in a human or animal host. antigens may be part of the physical structure of the pathogen, such as the bacterial cell wall, or they may be a chemical produced and released by the pathogen, such as an enzyme or toxin each antigen is also called an epitope.

antibodies: 

are globulin proteins (immune globulins) that react specifically with the antigen that stimulated their production. they make up about 20% of the protein in blood plasma. there are five classes of antibodies: igg, igm, iga,  igd and ige.

function of antibodies: 

neutralize toxins.  -

opsonize microbes. -

activate component.  -

prevent the attachment of microbes to mucosal surfaces. -

types of diagnostic tests: 

1. agglutination test:   

this reaction can be done in a small cup or tube or with a dropinginging on a slide.

one very commonly used agglutination test is the test that determines  a person’s abo blood group. 

abo blood groups: 

all human erythrocytes contain alloantigens of the abo group. there are four combinations of the a and b antigens called a, b, ab, o. 

a person’s blood group is determined by mixing the person’s blood with antiserum against a antigen on one area on a slide and with antiserum against b antigen on another area. if agglutination occurs only with a antiserum, the blood group is a if it occurs only with b antiserum, the blood group is b, if it occurs with both a and b antisera, the blood group is ab, and if it occurs with neither a nor b antisera, the blood group is o. 

2. complement fixation test: 

the reaction consists of the following two steps:

antigen and antibody (one known and the other unknown) are mixed, and a measured amount of complement (usually form guinea pig) is added. if the antigen and antibody match, they will combine and use up “fix” the complement. 

an indicator system, consisting of sensitized red blood cells (ex. red blood cells plus anti-red blood cells antibody), is added. if the antibody matched the antigen in the first step, complement was fixed and less (or non) is available to attach to the sensitized red blood cells. the red blood cells remain unhemolyzed, the test is positive because the patients serum had antibodies to that antigen. if the antibody did not match the antigen in the first step, complement is free to attach to the sensitized red blood cells and they are lysed, the test is negative. patient’s serum must be heated to (56°c) for 30 minutes to inactivate any human complement activity.   

3. precipitation test:

this test consist of the following:

precipitation is solution. 

precipitation in agar. 

precipitation in agar with an electric field. 

precipitation in agar: 

this is done as either single or double diffusion. 

1- single diffusion: 

antibody is incorporated into agar and antigen is measured into a well. as the antigen diffuses with time, precipitation rings form depending on the antigen concentration. 

ex. radial immunodiffusion is used to measure igg, igm, complements and other substances in serum. 

2- double diffusion: 

antigen and antibody are placed in different wells in agar and allowed to diffuse and form concentration gradients. where optimal proportions occur, lines of precipitate form. this method indicates whether antigens are identical, related but not identical, or not related.   

4. widal test: 

is a presumptive serological test for enteric fever or undulant fever. in case of salmonella infections, it is a demonstration of agglutinating antibodies against antigen o-somatic and h-flagellar in the blood. the test is not a very specific test, since patients are often exposed to other bacteria (ex. slamonella enteritidis, s. typhimurium and some types of e. coli).

procedure: 

patient’s serum is doubly diluted by mixing and transferring from 1:10 to 1:640 in three-four rows. first row usually comprises of felix tubes, where somatic s. typhi o antigen is added.

for all the remaining rows, dreyer’s tubes are taken where different flagellar h antigens are added. each tube must contain (0.5 ml) of diluted serum. a test tube with only saline is kept in each row as control. all the tubes (including control) in a row are mixed with (0.5 ml) of antigen suspension. the first row is treated with s. typhi o antigen, the second row with s. typhi h antigen, the third row with s. paratyphi ah antigen and the fourth row with s. paratyphi bh antigen. since infections by s. paratyphi b are rare, this antigen is usually omitted in the test. after all the tubes have been treated with specific antigen suspensions, the widal rack is placed in a thermostatically controlled water bath maintained at 37°c for overnight incubation. another approach is to incubate the tubes at 50-55°c. 

reading the results: 

the control tubes must be examined first, where they should give no agglutination. the agglutination of o antigen appears as a “math” or “carpet” at the bottom. agglutination of h antigens appears loose, wooly or cottony. the highest dilution of serum that produces a positive agglutination is taken as titre. the titres for all the antigens are noted.