The Western Blot – a tried and true experimental protocol where protein structures are separated via molecular weight/charge and transferred to a membrane before visualization by a chemiluminescent solution (say that three times fast!). Seems simple, right? While the step-by-step process of a western blot has for the most part remained the same over the years, variations in solutions, procedures and reagents may increase the efficacy of your results.
Western blotting is one of the most commonly used antibody assay techniques in cell and molecular biology research since its development over three decades ago, and is considered the gold standard for protein detection and quantification.
I first tried the PBP antibody (NB110-93495) in June of 2010 and it worked well. I picked this antibody because it had been tested in rat tissue, so I was confident it would work for my rat samples. I stored the PBP antibody in 20ul aliquots after it arrived then stored it at -20C and used it over a 3 month period. I ran a Western blot with 15ug of a RIPA whole cell lystate from WKPT cells a rat kidney immortalized cell line derived from the S1 proximal tubule segment.
The loading controls on our antibody database are widely used in gel electrophoresis and Western blotting studies. Products like the GAPDH antibody detect "housekeeping" proteins which are abundantly distributed in cells. This makes them useful for checking the even loading of gel samples, and the even transfer of proteins at the blotting stage. They also serve a purpose in quality control, by verifying reagents are working correctly, and in the standardization of experimental results.
Our antibody databaseincludes many thousands of proteins, and it is constantly being enriched. Modern developments mean that, whereas scientists would once have searched for one protein in a single sample, now they search for several – often simultaneously, and in minute quantities.
The vast majority of antibodies in our antibody catalog are suitable for Western blotting studies. Devised almost 30 years ago by W. Neal Burnette, it has become a standard assay wherever antibodies are used to detect proteins.
