with enzyme is added. The substrate specific to enzyme is added to well, and colored
product formed is measured. The sandwich ELISA is useful to detect antigen
concentration.
38.3 Competitive ELISA................................
Competitive ELISA is also used to measure antigen concentration. The process
involves the incubation of antibody in a sample solution containing antigen. This
antigen-antibody mixture is then transferred to a microplate well coated with anti-
gen. The assay is based on the fact that the presence of more number of antigens in
the sample will bind with more number of antibodies, and hence available free
antibody concentration that can bind with the antigen-coated well will be less. The
concentration of primary antibody that binds with antigen in the well can be detected
by addition of enzyme-conjugated secondary antibody specific for the isotype of the
primary antibody. The absorbance for this reaction will be inversely proportional to
the concentration of antigen in the original sample.
Fig. 38.1 Principle of
ELISA
154 38 To Perform Enzyme-Linked Immunosorbent Assay