This document describes detailed information about Radio immuno assay (RIA) including its principle, procedure, advantages, disadvantages, application etc
This document describes detailed information about Radio immuno assay (RIA) including its principle, procedure, advantages, disadvantages, application etc
Enzyme immunoassays (EIAs), also known as enzyme-linked immunosorbent assays (ELISAs), combine antibody binding with enzymatic detection to quantify molecules of interest.
A RIA is a very sensitive in vitro assay technique used to measure concentrations of substances, usually measuring antigen concentrations (for example, hormone levels in blood) by use of antibodies.
Immunodiffusion -Different Types,Principle,procedureand application. it is a diagnostic technique for the detection or measurements of antibodies and antigens by their precipitation which involves diffusion through a substances such as agar or gel agarose .common types -oudin procedure,oakley fulthorpe procedure ,mancini technique ,ouchterlony double immuno diffusion
In this slide contains types, working principle, factors affecting, advantage and disadvantage of paper electrophoresis.
Presented by: G.Sai Swetha. (Department of pharmacology),
RIPER, anantapur.
Enzyme immunoassays (EIAs), also known as enzyme-linked immunosorbent assays (ELISAs), combine antibody binding with enzymatic detection to quantify molecules of interest.
A RIA is a very sensitive in vitro assay technique used to measure concentrations of substances, usually measuring antigen concentrations (for example, hormone levels in blood) by use of antibodies.
Immunodiffusion -Different Types,Principle,procedureand application. it is a diagnostic technique for the detection or measurements of antibodies and antigens by their precipitation which involves diffusion through a substances such as agar or gel agarose .common types -oudin procedure,oakley fulthorpe procedure ,mancini technique ,ouchterlony double immuno diffusion
In this slide contains types, working principle, factors affecting, advantage and disadvantage of paper electrophoresis.
Presented by: G.Sai Swetha. (Department of pharmacology),
RIPER, anantapur.
The following presentation contains helpful information regarding Radioimmunoassay (RIA) and Enzyme-Linked Immunosorbent Assay (ELISA), including their history, introduction, advantages, procedures and applications.
Radioimmunoassay (RIA) is a very sensitive in vitro assay technique used to measure concentrations of antigens (for example, hormone levels in blood) by use of antibodies. As such, it can be seen as the inverse of a radiobinding assay, which quantifies an antibody by use of corresponding antigens.
Immunoassay is a biochemical test that estimate or asses the presence or concentration of a macromolecule (antigen) in a solution (eg-blood) through the use of an antibody or immunoglobulin(Ig). The macromolecule called "analyte". Analytes in biological liquids such as blooed serum, biological fuid and urine are frequently measured using immunoassays ( for medical and research purposes).
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3. INTRODUCTION
• Radio immuno assay is an immunological assay to
analyse antigens present in given biological samples
• It is most sensitive and specific method of immuno
assay
• Sensitivity ranges from 0.0006 -0.006 µg / ml .
• It was developed by S.A. Berson and Rosalyn yalow
in 1959 in received nobel prize in 1977.
4. THEORY
• RIA involves the reaction between antigen (hapten) and its specific
Antibody in presence of radio labelled antigen.
• This hapten is obtained from a non-antigenic compound e.g.,
morphine , Cartelol etc., which is ultimately conjugated .
• Animals normally develop antibodies to the injected immunogenic
substances as part of their natural immune response.
• The serum derived from this animals is used as the antibody source
and tested with reference to their specificity, affinity at their titre
level.
5. PRINCIPLE
• It involves three principles which make it most specific & sensitive than
other immune assays.
• An immune reaction i.e. antigen, antibody binding.
• A competitive binding or competitive displacement
reaction (It gives specificity).
• Measurement of radio emission (It gives sensitivity).
6.
7. REQUIREMENTS
• Micro titer plate / Test tubes.
• Pure antigen
• Radio labelled antigen
• Antibodies
• Standard's
• Centrifuge
• Radioactive counter
8. Micro titer plate :
• A microtiter plate is used mostly used for this assay
• A microtiter plate could have 6, 24, 96, 384 or even sometimes
1536 wells arranged in rows.
• Each well of a microplate can only hold very small amounts of
liquid.
9. PURE ANTIGEN :
• Antigen may be obtained from biological sample or by synthetic form , it
should be pure.
• It is used as standard or calibrator.
RADIO LABELLING OF ANTIGEN:
•The most commonly used radiolabels in RIA are tritium and iodine
• They have adequate activity and have long enough
half lifes.
10. ANTIBODY
• Specific antibodies are obtained by injecting the
Ag to animals.
• Ag I.e., drug molecule + bovineserum albumin
12. Centrifuge:
• Used foe the separation of
precipitated form and
supernatant liquid form.
• Range: 1200 – 2500 rpm
13. Radio active counters :
Two types of counters are used
They are :
a) Gamma counter.
b) Scintillation counters.
14. a)Gamma counter :
• These are used for the gamma energy emitting isotopes
e.g., common iodine isotopes.
b) Scintillation counter :
• These are used for counting beta energy emitting isotopes
e.g., tritium , carbon -14 isotopes
16. Test tube-1
Blank
Ab +Ag*
Test tube -2
Calibrator
Ab + pure Ag +Ag*
Test tube -3
Sample
Ab + sample Ag + Ag*
PROCEDURE :
17. SEPARATION TECHNIQUES :
• After completion of reaction of reaction free form and bound forms
are determined by separation techniques.
• Various techniques include gel filtration, Electrophoresis, solid
phase adsorption of Ag, Ab & fractional precipitate.
18. Applications :
Radio immuno assay of morphine :
This involves :
• Synthesis of immunogen
• Antiserum production.
• Procedure.
19.
20. CONCLUSION :
• RIA is invitro assay technique used for the determination of Ag concentration
present in given sample.
• In advance to RIA many other techniques are introduced like RAST, EIA , RIST,
lRMA , competitive RIA.