Eastern Blotting - History and Multiple Definitions

History and Multiple Definitions

. The current definitions are summarized below in order of the first use of the name; however, all are based on some earlier works. In some cases, the technique had been in practice for some time before the introduction of the term.

• (1982) The term eastern blotting was specifically rejected by two separate groups: Reinhart and Malamud referred to a protein blot of a native gel as a native blot; Peferoen et al., opted to refer to their method of drawing SDS-gel separated proteins onto nitrocellulose using a vacuum as Vacuum blotting.
• (1984) Middle eastern blotting has been described as a blot of polyA RNA (resolved by agarose) which is then immobilized. The immobilized RNA is then probed using DNA.
• (1996) Eastern-Western blot was first used by Bogdanov et al. The method involved blotting of phospholipids on PVDF or nitrocellulose membrane prior to transfer of proteins onto the same nitrocellulose membrane by conventional Western blotting and probing with conformation specific antibodies. This method is based on earlier work by Taki et al. in 1994, which they originally dubbed TLC blotting, and was based on a similar method introduced by Towbin in 1984.
• (2000) Far-Eastern blotting seems to have been first named in 2000 by Ishikawa & Taki. The method is described more fully in the article on Far-eastern blotting, but is based on antibody or lectin staining of lipids transferred to PVDF membranes.
• (2001) Eastern blotting was described as a technique for detecting glycoconjugates generated by blotting BSA onto PVDF membranes, followed by periodate treatment. The oxidized protein is then treated with a complex mixture, generating a new conjugate on the membrane. The membrane is then probed with antibodies for epitopes of interest. This method has also been discussed in later work by the same group. The method is essentially Far-Eastern blotting.
• (2002) Eastern blot has also been used to describe an immunoblot performed on proteins blotted to a PVDF membrane from a PAGE gel run with opposite polarity. Since this is essentially a Western blot, the charge reversal was used to dub this method an Eastern blot.
• (2005) Eastern blot has been used to describe a blot of proteins on PVDF membrane where the probe is an aptamer rather than an antibody. This could be seen as similar to a Southern blot, however the interaction is between a DNA molecule(the aptamer) and a protein, rather than two DNA molecules. The method is similar to South-western blotting.
• (2006) Eastern blotting has been used to refer to the detection of fusion proteins through complementation. The name is based on the use of an enzyme activator (EA) as part of the detection.
• (2009) Eastern blotting has most recently been re-dubbed by Thomas et al. as a technique which probes proteins blotted to PVDF membrane with lectins, cholera toxin and chemical stains to detect glycosylated, lipoylated or phosphorylated proteins. These authors distinguish the method from the Far-eastern blot named by Taki et al. in that they use lectin probes and other staining reagents.
• (2009) Eastern blot has been used to describe a blot of proteins on nitrocellulose membrane where the probe is an aptamer rather than an antibody. The method is similar to South-western blotting.

There is clearly no single accepted definition of the term. A recent highlight article has interviewed Ed Southern, originator of the Southern blot, regarding a re-christening of Eastern blotting from Tanaka et al. The article likens the Eastern blot to "fairies, unicorns, and a free lunch" and states that Eastern blots "don't exist." The Eastern blot is mentioned in an Immunology textbook which compares the common blotting methods (Southern, Northern, and Western), and states that "the Eastern blot, however, exists only in test questions."

The principles used for Eastern blotting to detect glycans can be traced back to the use of lectins to detect protein glycosylation. The earliest example for this mode of detection is Tanner and Anstee in 1976, where lectins were used to detect glycosylated proteins isolated from human erythrocytes. The specific detection of glycosylation through blotting is usually referred to as lectin blotting. A summary of more recent improvements of the protocol has been provided by H. Freeze.