I am doing the rsearch in cardiovascular system. In my project, I am trying to qunatify the protein content from mice carotid artery. My problem is, After quanitfication, when I ran the gel, I didnt get the equal band size for all the sample ( with commasiev staining as well as in the Western blot)...
I have used the 2x SDS buffer for lysis the tissue and then centrifuged @ 14000 rpm for 15 min. After that I took the supernatant for the protein qunatification. I did the protein quantification with BCA protein assay then based on the value I have loaded 10 microgram (Protein) of sample to each well.
I have tried various steps and repeated 3-4 times ... I feel there is a problem in lysis buffer...!!!! I want to know the buffer which I am using (2 x SDS) is correct or not.....!!! I need some idea for you to solve this problem.