Elisa
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The document discusses enzyme-linked immunosorbent assay (ELISA), including its introduction, principle, equipment, procedure, types, advantages, and disadvantages. ELISA is a qualitative or quantitative immunological procedure that detects antigens or antibodies using enzyme-labeled antibodies and chromogenic substrates. It relies on antibody-antigen interactions and uses an enzyme-labeled antibody to generate a colored reaction, allowing detection of a particular antigen. The document outlines the basic equipment, general procedure involving coating wells with antibodies and adding samples and enzyme-labeled antibodies, and the three main types of ELISA - indirect, sandwich, and competitive.
Elisa
- 1. BARKATULLAH UNIVERSITY, BHOPAL ENZYME LINKED IMMUNOSORBENT ESSAY (ELISA) Presented by: Navjot Singh M.Sc. 3rd semester Enrollment Number – R218145050002 Roll Number – 2181400013
- 2. INTRODUCTION ELISA is a qualitative or quantitative immunological procedures in which the Ag-Ab reaction is monitored by enzyme measurements. The term ELISA was first used by Engvall & Perlma in 1971. The ELISA test was the first screening test commonly employed for HIV. It has a high sensitivity.
- 3. Principle Its principle is similar to RIA but depends on an enzyme rather than a radioactive label. An enzyme conjugated with an antibody reacts with a colorless substrate to generate a colored reaction product. Such a substrate is called a chromogenic substrate. A number of enzymes have been employed for ELISA including alkaline phosphatase, horseradish peroxidase and β-galactosidase.
- 4. Equipment's Microwell plate It is flat bottom polystyrene plate containing 8 x 12 wells holding 350 µl each.
- 5. Equipment's Multi-pipette It is an 8-channel 100 μL pipette which is a good help for even small scale work.
- 6. Equipment's Washing device It is a manually operated washing device.
- 7. Equipment's Microplate washer These are very efficient with very low carry over contamination.
- 8. General Procedure of ELISA The wells are coated with antibodies. Sample is added which may contain antigen. The antigen-antibody interaction took place. Removal of unbound antigens by washing. Addition of another antibody which is linked with enzyme. Interaction of enzyme with particular substrate. This interaction produces color through which we can observe a particular antigen (disease).
- 9. Types of ELISA There are three types of ELISA :- Indirect ELISA Sandwich ELISA Competitive ELISA
- 10. Types of ELISA Indirect ELISA
- 11. Types of ELISA Sandwich ELISA
- 12. Types of ELISA Competitive ELISA
- 13. Advantages of ELISA Reagents are relatively cheap and have a long shelf life. ELISA is highly specific and sensitive. No radiation hazards occur during labelling or disposal of waste. Easy to perform and quick procedures. Equipment's are inexpensive and widely available. ELISA can be used to detect variety of infections.
- 14. Disadvantages of ELISA Measurement of enzyme can be more complex. Enzyme activity may be affected by plasma constituents. Kits are commercially available, but not cheap. Very specific to a particular antigen. Won’t recognize any other antigen. False positives/negatives possible, especially with mutated/ altered antigen.
- 15. References Kuby J., Osborne B.A, Goldsby R.A, “Immunology”, 5th edition. Pg. no 148-150 Ochei, J. and Kolhatkar A., (2015), Medical Laboratory Science, Theory and Practices, Tata McGraw-Hill.
- 16. Thank You