ELISA is a biochemical technique used in immunology to detect the presence of an antibody or antigen in a sample. It involves coating microtiter plate wells with an antigen or antibody and using conjugated enzymes and substrates to produce a colored product to indicate a positive result. There are different types of ELISA including direct, indirect, sandwich, and competitive ELISA which are used to test for various antigens or antibodies. ELISA has many applications such as measuring serum antibody concentrations, detecting food allergens or diseases, and identifying past exposure to diseases.

1. ELISA TO Ms.Sannia By Urooj Sabar 4318 5 th Semester (Morning) Bioinformatics & Biotechnology

2. Enzyme-Linked Immuno sorbent Assay INTRODUCTION Is a biochemical technique used mainly in immunology to detect the presence of an antibody or an antigen in a sample. first and most basic test to determine if an individual is positive for a selected pathogen, such as HIV.

3. 8 cm x 12 cm plastic plate which contains an 8 x 12 matrix of 96 wells, each of which are about 1 cm high and 0.7 cm in diameter.

4. What To Detect or Measure? ELISA provide a useful measurement of antigen or antibody concentration. detect the presence of antigens that are recognized by an antibody test antibodies that recognize an antigen.

5. An ELISA is a five-step procedure Coat the micro titer plate wells with antigen or Antibody.

6. Second step block all unbound sites to prevent false positive results using the blocking solution.

7. 3 rd step Add sample to the wells.

8. add anti-mouse IgG (antibody) conjugated to an enzyme. enzyme – alkaline phosphatase is used commonly 4 th step

9. 5 th step reaction of a substrate with the enzyme to produce a colored product, thus indicating a positive reaction.

10. Types Of ELISA Direct ELISA Indirect ELISA Sandwich ELISA Competitive ELISA

11. Direct ELISA Used to test the presence of a specific antigen in sample.

12. Indirect ELISA Used to test the presence of a specific antibody in a sample.

13. Direct Sandwich ELISA

14. Indirect Sandwich ELISA

15. Competitive ELISA The labelled antigen competes for primary antibody binding sites with the sample antigen (unlabeled). The more antigen in the sample, the less labelled antigen is retained in the well and the weaker the signal).

17. Applications Serum Antibody Concentrations Detecting potential food allergens (milk, peanuts, walnuts, almonds and eggs) Disease outbreaks- tracking the spread of disease e.g. HIV, bird flu, common, colds, cholera, STD etc Detections of antigens e.g. pregnancy hormones, drug allergen, GMO, mad cow disease Detection of antibodies in blood sample for past exposure to disease e.g. Lyme Disease, trichinosis, HIV, bird flu

18. Any Question?