Anti-Cow Serum Lab Report

Introduction:
The purpose of this experiment is to see the anti-cow antibody bind to cow serum only, and we expect to see the anti-cow antibody bind to the spot that had the cow serum. The system we use in this experiment is the serum from Cow, Horse, Goat, Sheep, and Donkey, Chicken. In order to able to detect and analyze proteins based on their ability to bind to a specific antibody, the SDS-PAGE and Western Blot was performed.
SDS Polyacrylamide Gel Electrophoresis (SDS-PAGE) is a very common technique used to separate proteins based on molecular weight under the influence of an applied electrical field and then used to prepare for the Western Blot (#1 Lehninger). The support medium used is a polyacrylamide gel and, it also used sodium
The stacking gel was used to make sure all the proteins start separating at about the same time. It has a larger pore size so that the larger protein can move in easily. The SDS and heating was used to denature the proteins, the proteins then loaded into the lane. When apply the electrical field, the proteins will move toward the bottom, which is the positive anode. Also, the Tris-glycine will enter the stacking gel. The pH of stacking gel is 6.8, it makes the proteins less negative because the amino acids are protonated at equilibrium. Because the glycine moves slower than chloride ion, a Gly-chloride ion boundary is formed. But, glycine still runs little bit faster than other proteins. Therefore, the proteins are trapped between chloride ion and glycine. A very tight band is formed inside the stacking gel by the
It was increase from 6.8 to 8.8 and the pores are also smaller. Due to the pH different, the N-terminal amino groups are deprotonated. At the equilibrium the proteins are more negatively charged compared when it in the stacking gel. Therefore, glycine moves faster than proteins. Due to the smaller pores size in the resolving gel, the smaller the protein will move faster toward the positive anode or the bottom (#2 ruf.rice.edu). Western Blot is a common used technique to identify and analyze proteins according to their ability to bind to a specific antibody. It is an analytical method that protein sample was first separate based on the molecular weight using the SDS- PAGE method, and then transferred on the nitrocellulose. The specific primary enzymes labeled antibody was used to detect the transferred protein. Antibodies bind to specific sequences of amino acids, and can recognize specific proteins among a group of many because the amino acid sequences are different from protein to protein (#3