Hi Everyone,
I used HDAC9 monoclonal antibody from a company which is suppose to be 120kDa. However really clear and bulk of band (only band in gel) appear at around 55 kDa, I realized that this company also had HDAC9 recombinant human protein at around 50.7kDa. In literature, there is not a consistent data provided. How come any protein is available commercially in many versions and it is so debatable?
Thank you
HDAC9 has 11 isoforms. The antibody you're using recognizes one or many of them...
Here are the isoforms: https://www.uniprot.org/uniprot/Q9UKV0
Here's Abcam antibody. the band is around the same size as yours - the Ab recognizes isoforms 3, 8, 9, 10, and 11. link: https://www.abcam.com/hdac9-antibody-ab59718.html
Careful what Ab companies claim...in their mind, quality control is typically incumbent upon the user. They might have used a shorter, recombinant form for both Ab generation and Western validation. According to uniprot (fairly accurate usually, for the canonical form anyways) HDAC9 is around 100 kDa (major form). The most likely reason why you're not picking up a 100 kda signal is that protein expression is too low. Do you have any expression data (mRNA) showing expression in your system, and if so, is it high? You may find the info at Gtex or https://www.proteinatlas.org.
See also: https://www.ncbi.nlm.nih.gov/gene?cmd=Retrieve&dopt=Graphics&list_uids=9734
According to source above, expression fairly low in primary tissues. May be different in a cell model however (protein atlas suggests a fairly high nuclear expression in immortalized cells).
As for the 55 kda, it is probably an (abundant) cross-reacting band.
If you're embarking on a HDAC9 project, I would definitely advise you perform an orthogonal validation (siRNA, KO etc...) of HDAC9 in your system first.
Good luck
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