I often think that we, young scientists resemble to kids with cell phones or tablets in the way that they we rely on technology and learn it fast but don’t really understand what comes behind the creation of such devices. It is amusing by how the passage of time erases memory of the hard work that goes into developing ‘common use’ or new biochemical tools. That's why I'll write a little about the so commonly used and usefull Western Blot.
The name of this technique was given by W. Neal Burnette to keep the compass theme for naming blotting methods (Northern and Southern) and because his lab was located on the West Coast
Now, Western blotting was developed by three groups almost simultaneously and of course with some differences and so the credits should go to ALL of them: George Starck’s, Harry Towbin’s and W. Neal Burnette’s.
The Stark’s group was already famous for developing the RNA blotting technique called “Northern blot”, a name that came as a joke based on the DNA blotting technique called “Southern blot,” which in turn was named after its inventor, Edwin Southern. Stark’s lab became experts in making diazobenzyloxymethyl-paper or DBM-paper, which they use for Northern blots. They soon realized that there was a problem in detecting specific proteins and used the same chemistry as for Northern blotting, because they realized DBM-paper reacted with both nucleic acids and proteins. When the proteins transferred out of the gel, they covalently bound to the DBM-paper. “The surprise bit was that the immobilized protein reacted so nicely and specifically with antibodies” notes Stark (1). The Stark group published a technique for passively transferring proteins to DBM-paper which were then detected by radio-labeled Protein A.
Meanwhile, Towbin, a postdoc in Switzerland was focusing on making antibodies against ribosomes for structure-function studies for his research project, but didn’t have a biochemical tool for the analysis. So he began to work out a method that would allow him to establish which antibody bound to which component of the ribosomal complex. By that point, DNA and RNA blotting methods were popular, so the idea of transferring proteins out of a gel and onto a membrane seemed natural. “It was in the air!” says Towbin (1). He says they knew that proteins, but not RNA, bound to nitrocellulose, so they separated ribosomal proteins on a polyacrylamide gel with urea as a denaturing agent and then electrophoretically transferred them onto nitrocellulose. Although they primarily focused on gels with urea as the denaturing agent, Towbin says they also got their approach to work with SDS. The Towbin group published a technique that is very similar to the one used today. Their technique used electric current to transfer proteins to nitrocellulose membranes within the standard “immunoblotting sandwich.” Additionally, they used secondary antibodies to detect bound proteins.
Burnette was unaware of the work done by the two groups as he was developing his approach but saw the Stark and Gordon groups’ papers in print while he was preparing his manuscript. But he felt that his method was different enough to press ahead. He focused on electrophoretically transferring proteins out of SDS-polyacrylamide gels onto nitrocellulose in a more quantitative manner. The Burnette group published an electrophoretically method of protein transfer to nitrocellulose which were then detected by radio-labeled Protein A.
The Starck and Towbin groups published almost simultaneously in 1979. The Burnette group, although submitting their paper that same year, it was rejected and published later on in 1981. However, Burnette is often cited as the developer of the western blot probably because he used a good marketing technique by choosing a clever name. Funny thing is that Burnette himself cites this naming of the technique as one of the reason his paper was rejected, since a reviewer said it was given a ‘flippant name’.
Interesting how a ‘common use’ technique involved so much work. Towbin once said “The younger generation of biologists takes the method for granted!”.
My advice: do not spend all of your time studying for exams and take some time to learn about the history and the people involved in the bench tools you use on a daily basis, take time to really understand what you are doing and how things work and be thankful that someone did an amazing job so you can do yours today.
1. http://www.asbmb.org/asbmbtoday/asbmbtoday_article.aspx?id=16084
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