Question

11. What substance did we use as negative control in our ELISA experiment? (2 pts) 12. We rinsed the wells with twice with PBS-Tween and the only once with PBS afterwards. What is the purpose of rinsing with PBS after PBS-Tween? (5 pts) 13. Name one infectious disease that is usually diagnosed by an Indirect ELISA method: (3 pts) 14. What substance did we used as a blocking agent in the ELISA experiment? (5 pts) A. PBS-Tween B. Biotin C. Streptavidin-peroxidase D. BSA E. TMB 15. We put "blocking reagent" after coating the wells with antigen. What is the purpose of that? Explain what may happen if you don't put the blocking reagent in. (5 pts) 16. During electrophoresis, the gas that comes out at the positive end is: (3 pts)

Answer 1

11.Distilled water is usually used as negative control. It should not contain any primary antibody.

12.PBS buffer is added as it maintains isotonicity.Tween is a detergent used in washing buffer. So sometimes, it may cause froth. Hence, again the wells are washed with PBS.It prevents non specific binding of proteins.

13.Swine flu ,Toxoplasmicosis are few of the infectious diseases detected through indirect elisa.

14.BSA is usually used as a blocking agent in ELISA.

15.Blocking agents are those that does not have binding affinity for the target.To ensure specificity and eliminate residual binding capacity, blocking agents are used.They reduce non specific interactions and stabilize biomolecules bound the the surface of the wells.

16.Oxygen gas is released at the positive end, which immediately form hydroxonium ions with wate molecules.

17.Hydrogen gas is released at the negative end, due to the reduction reaction with water.

19.The restriction enzyme cuts at 4 positions as given in the problem.So there will be 5 fragments .The fragments are cut at 1500 position, next 2700 position.So the weight of that fragment would be 2700-1500=1200bp.Similarly, rest follows. The weight of the fragments would be 1500 bp, 1200bp, 900 bp(3600-2700) ,400bp and 1000 bp. Totally 5000bp.