Explain with examples the use of antibodies in bio or immuno assays.

Immunoassay
Every immunoassay system relies on immunological reagents as its foundation. Intricate amounts of
analytes in complicated biological systems may be quantitatively and qualitatively measured using
immunoassays. Although the ideas underpinning the creation of an immune test appear simple, it is
typical for assays to be created that give inaccurate or misleading results when the right selection of
immunoreagents is not a crucial component of the assay design. There are several different supports and
detecting methods used in immunoassays. The amount of sensitivity and specificity of each immunoassay
is, however, controlled by an antibody at its core.

Different types of Immunoassays

Immunoassays are tests that use antibody-antigen binding interactions to find out whether a certain
molecule is present in a sample. Immunoassays are very specific because antibodies only attach to a
certain structure of a given antigen, which makes antibodies unique to that structure. Because of this,
antibodies are useful tools for identifying target molecules. Immunoassays are therefore a crucial tool for
medical facilities, life science research, and business laboratories. Immunoassays are available in a
variety of forms and may be used to trace proteins, evaluate illness, and find environmental contaminants.
The four most commonly used types of labeled immunoassays are:

1. Enzyme Immunoassay (EIA) or Enzyme-linked immunosorbent assay (ELISA)

2. Radioimmunoassay (RIA)
3. Fluoroimmnoassay (FIA)
4. Chemiluminescence immunoassay (CLIA)
 
 Enzyme-linked immunosorbent assay
Simple, frequently used tests called enzyme immunoassays, or ELISAs, rely on extremely precise
antibody-antigen interactions. An ELISA is made to identify and measure soluble substances
including hormones, proteins, and antibodies. Sandwich, competitive, direct, and indirect tests are
the four primary ELISA assay types. Both the way the antibody or antigen is bound to the solid
plate and how the signal is measured varies across these procedures.

 Radioimmunoassay

A radioimmunoassay (RIA) measures the amounts of chemicals in bodily fluids including blood
and saliva using radiolabeled antigens. An interesting antigen is associated with a radioisotope by
its corresponding antibody. The radioactive antigen is subsequently replaced at the binding site
with a sample containing the target antigen, which competes with it.

After unbound antigens have been removed by washing, the sample's radioactivity is assessed.
More target antigen is present when the radioactive signal is less. Due to the radioactive materials
 used, more safety measures are required compared to other immunoassay procedures, yet RIA's
excellent sensitivity and specificity ensure that it is still in use today.

 Fluoroimmunoassay
Various chemicals can be detected and quantified using fluorescent immunoassays (FIA), which
employ a fluorescent compound as the detection reagent. In comparison to other techniques, FIA is
popular in the in vitro diagnostics (IVD) market because it is quick and extremely sensitive.
Fluorescent probes are used to identify antibodies in FIA. UV-illuminating FIA fluorescent dyes
are employed to identify a particular antigen-antibody binding. The antibody-antigen complexes
are separated after being exposed to the antigen, and the fluorescence intensity is then calculated.

 Chemiluminescence immunoassay
An ELISA or fluoroimmunoassay uses the same basic concepts as a chemiluminescent
immunoassay (CLIA), but uses a different reporter. When an electron is pushed to a higher energy
level and then relaxes down, it releases light, which is known as luminescence. FIA employs
certain light frequencies, whereas CLIA uses a chemical process to boost the electron's energy.
CLIA is employed in a variety of industries due to its high sensitivity and specificity, including
environmental monitoring, illness diagnostics, life sciences, and food safety. Additionally, newer
methods like magnetic-bead based CLIA have increased its potential. For instance,
chemiluminescence tests activated by magnets were created to find the Zika virus in patient
samples.

Application of Antibodies in Immunoassays

The antibodies can be then used in a multitude of applications, including but not limited to: western blot
(WB), immunoprecipitation (IP), immunofluorescence (IF), immunohistochemistry (IHC), chromatin
immunoprecipitation (ChIP), and flow cytometry (FC). As a cornerstone of the body’s immune response,
antibodies can provide significant data to support scientists’ research. In immunological studies,
antibodies can be used in vivo to deplete specific cells for functional analyses. Antibodies are also in vivo
for neutralization of cell surface receptors to enable binding to soluble factors.