Question

Question 12 parts a,b,c

Q12. Using the information provided below on ELISA, answer the following questions

Answer 1

ELISA is a technique which uses concept of antigen antibody interaction to identify and quantify expression pattern of a protein. A primary antibody identifies a epitope (part of protein- here antigen which attaches with antibody) in the protein of interest and binds to it. Then a secondary antibody is used for which primary antibody is antigen then secondary antibody linked with enzyme attaches itself to primary antibody. Now substrate for the enzyme is added. More the attached primary antibody- more will be attched secondary antibody hence more enzyme thus intense colour will appear due to chromogenic substrate enzyme interaction. This technique helps in checking relative expression of any protein. Less intense colour means less expression of protein and vice versa.

While designing any experiment we take two controls i.e. positive control (pure chip28 protein) and Negative control ( Only buffer which we have used to isolate chip 28 from different organs but without any cell). Tissue extract of different organs will be prepared by using buffer. Now we will be adding equal volume (Suppose 25ul) of all the extract in different veils of microtitter plate. Then we will add Primary/secondary antibody then substrate for the enzyme. The highest colour will appear in the positive control i.e. pire chip 88 and No colour will develop in Negative control veil. All other viels will show intensity somewherre in between both the controls. The ccell extraxt showing closest intensity to positive control veil will be highest expresssing cell.

Aquaporins are also known as water channels, through which water can flow more rapidly into and out of the cell than by diffusing through the phospholipid bilayer . The proximal tubules of kidney (Nephrons) are extensively involved in absorption of water so it is only logical that it expresses chip 88 in significantly high amount.