This document discusses immunoassays and two common types - radioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA). RIA was developed in the 1950s and involves competition between labeled and unlabeled antigens for binding with antibodies. Radioactivity of the antigen-antibody complex is then measured. ELISA uses an enzyme like alkaline phosphate to visualize antigen-antibody reactions. It has two types - direct ELISA using a double antibody sandwich and indirect ELISA using direct antigen coating. Both assays are used to detect various antigens and antibodies and have applications in clinical diagnosis, food testing, and toxicology.

1. Presented By-
ROHIT
R.K.S.D college of
pharmacy, Kaithal (Hry)
M.Pharma 1st year
(Pharmaceutics)

2. An immunoassay is a biochemical test that
measures the presence or concentration of
a macromolecule or a small molecule in a solution
through the use of an antibody (usually) or
an antigen (sometimes).
The molecule detected by the immunoassay is
often referred to as an "analyte" and is in many
cases a protein.

3. IMMUNOLOGICAL ASSAY.
RIA
ELISA

4. 1.RIA (RADIO IMMUNO ASSAY)
It was developed by “Rosalyan Yalow and S.
A. Berson in the 1950s”.
Development of the ‘RIA for insulin;
DEFINATION –
It is a scientific method used to
test antigens without the need to use a
bioassay.

5. PRINCIPLE
It involve competition between
labelled and unlabelled antigen for binding with
specific antibodies so antigen-antibody
complex is formed .& radio activity is measured.

6. INSRUMENTATION
It is an alternative technique employed in
diagnosis of several antigen like hepatitis.
It is competitive binding assay in which serum
sample is combined with radioactively labelled
antigen and antibodies to the antigen .
The radioactive antigen and antigen specimen
will compete for antibody .the antigen-antibody
complex thus formed is isolated and analyzed for its
radioactivity. The concentrations ‘standard dose
response curves;

7. DIAGRAMMATIC REPRESENTATION OF RIA.

8. APPLICATION
1. Detection of insulin.
2. Detection of narcotic drug .(morphine and
heroin )
3. Detection of digoxin .
4. Determination hydromorphon and
hydrocondron in human serum .
5. Measurement of ferritin.
6. Thyroid testing .

9. 2.ELISA (ENZYME LINKED IMMUNOSORBENTASSAYS)
The ELISA is a fundamental tool of clinical
immunology and is used as an initial screen for
HIV detection.
Definition –
Enzyme-linked ImmunoSorbent Assay ,also
called ELISA ,enzyme Immuno Assay or ELA is
a biochemical technique used mainly in
immunology to detect the presence of an
antibody or an antigen in a sample.

10. PRINCIPLE
Antigen –antibody interaction this test
allows for easy visualization of results and can
be completed without the additional concern of
radioactive material used.

11. INSTRUMENTATION
In which enzyme like alkaline phosphate
is employed to visualize antigen-antibody
reaction.
TWO TYPE –
1. Direct ELISA ( double antibody sandwich)
2. Indirect ELISA ( direct antigen coating )

12. DIAGRAMMATIC REPRESENTATION OF ELISA.

13. 1. DIRECT ELISA
It is performed in microtitre
well, in which antibody to specific antigen is
adsorbed to well, on which specimen
containing antigen to be demonstrated is
added. Unbound antigen are washed off and
enzyme –linked antibody specific for test
antigen is added which binds to complexed
antigen forming sandwich.

14. 2. INDIRECT ELISA -
It is also performed in microtitre well, in
which antigen to the antibodies demonstrated is
adsorbed to well ,on to which patient antiserum
specimen is added If serum contains specific
antibody ,it will bind to the adsorbed antigen
.un-reacted serum is washed off and enzyme –
linked anti-HSIG is added, which will bind to
antibodies in the initially formed antigen
antibody complex.

15. APPLICATION
1. It is a useful tool for determination serum
antibody concentrations.( such as with the HIV
TEST)
2. For detecting the presence of antigen .it has
also found applications in the food industry in
detecting potential food allergens such as milk
,almond, eggs

16. APPLICATION
3. ELISA can also be used in toxicology as a rapid
presumptive screen for certain classes of drug .
4. Diagnosis of infection .