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Enzyme-linked immunosorbent assay (ELISA)

Enzyme-linked immunosorbent assay (ELISA). Lab 13. ELISA for Detection & Measurement of Antigen. 1. Add test sample to well of ELISA plate. 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. A. 2. Wash well to remove unbound antigens. B. C. D. E. F. 3. Add antibody-enzyme conjugate. G.

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Enzyme-linked immunosorbent assay (ELISA)

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  1. Enzyme-linked immunosorbent assay (ELISA) Lab 13

  2. ELISA for Detection & Measurement of Antigen 1. Add test sample to well of ELISA plate 1 2 3 4 5 6 7 8 9 10 11 12 A 2. Wash well to remove unbound antigens B C D E F 3. Add antibody-enzyme conjugate G H 4. Wash well to remove unbound conjugate 5. Add chromogenic substrate for enzyme Example - Feline leukaemia virus antigen (p27) test 6. Read absorbance on microplate reader

  3. ELISA for Detection & Measurement of Antibody 1. Well coated with antigen Examples Detection of collagen-specific antibody in canine SLE 2. Add test serum sample anti-dog Ig HRP autoantibody collagen 3. Wash well to remove unbound conjugate Detection of FIV-specific antibody 4. Add antibody-enzyme conjugate specific for the immunoglobulin in the test serum anti-cat Ig-HRP anti-FIV antibody 5. Wash well to remove unbound conjugate viral peptide 6. Add chromogenic substrate for enzyme 7. Read absorbance on microplate reader

  4. Immunoassay using labelled competitor 1. Add test sample (10ml) to well of ELISA plate wells are pre-coated with anti-progesterone antibody progesterone-enzyme complex 2. Progesterone in the sample binds to coating antibody concentration 3. Add progesterone-enzyme (alkaline phosphatase) conjugate 200ml progesterone in milk sample colour 4. Wash well to remove unbound progesterone-enzyme complex 5. Add chromogenic substrate for enzyme

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