Microbiology, 2021

20.4 • EIAs and ELISAs 831
Mechanisms of Select Antibody-Antigen Assays
Type of Assay Mechanism Example
Direct: Some bacteria and viruses cross-link red blood
cells and clump them together
Diagnosing influenza,
mumps, and measles
Direct Coombs’ test (DAT): Detects nonagglutinating
antibodies or complement proteins on red blood cells in
vivo
Checking for maternal
antibodies binding to
neonatal red blood cells
Hemagglutination
Indirect Coombs’ test (IAT): Screens an individual for
antibodies against red blood cell antigens (other than
the A and B antigens) that are unbound in a patient’s
serum in vitro
Performing pretransfusion
blood testing
Viral hemagglutination inhibition: Uses antibodies from
a patient to inhibit viral agglutination
Diagnosing various viral
diseases by the presence of
patient antibodies against
the virus
Blood typing and cross-matching: Detects ABO, Rh, and
minor antigens in the blood
Matches donor blood to
recipient immune
requirements
Table 20.3
20.4 EIAs and ELISAs
Learning Objectives
By the end of this section, you will be able to:
• Explain the differences and similarities between EIA, FEIA, and ELISA
• Describe the difference and similarities between immunohistochemistry and immunocytochemistry
• Describe the different purposes of direct and indirect ELISA
Similar to the western blot, enzyme immunoassays (EIAs) use antibodies to detect the presence of antigens.
However, EIAs differ from western blots in that the assays are conducted in microtiter plates or in vivo rather
than on an absorbent membrane. There are many different types of EIAs, but they all involve an antibody
molecule whose constant region binds an enzyme, leaving the variable region free to bind its specific antigen.
The addition of a substrate for the enzyme allows the antigen to be visualized or quantified (Figure 20.22).
In EIAs, the substrate for the enzyme is most often a chromogen, a colorless molecule that is converted into a
colored end product. The most widely used enzymes are alkaline phosphatase and horseradish peroxidase for
which appropriate substrates are readily available. In some EIAs, the substrate is a fluorogen, a nonfluorescent
molecule that the enzyme converts into a fluorescent form. EIAs that utilize a fluorogen are called fluorescent
enzyme immunoassays (FEIAs). Fluorescence can be detected by either a fluorescence microscope or a
spectrophotometer.