Five great application tips for proteomics research

October 31, 2011 at 10:44 am 2 comments

Here are some great application tips from Bio-Rad Laboratories for those researchers working with proteins:

  • Generally, the best method for keeping a protein in solution is to add any combination of nonionic detergents, zwitterionic detergents, and chaotropic agents to the sample mixture. Also use reducing agents such as DTT and DTE (less than 20 mM) to decrease disulfide bond formation between proteins.
  • When working with membrane or insoluble proteins, increase the amount of SDS in the equilibration and running buffers (up to 0.2%) to allow the proteins to effectively migrate out of the IPG strip.
  • To reduce the amount of SDS in samples generated by preparative SDS-PAGE, substitute the elution buffer with one that does not contain SDS.
  • Nucleic acid contamination is a common cause of horizontal gel streaking. Treat samples with nucleases to remove nucleic acids prior to isoelectric focusing.
  • Never heat samples in urea-containing buffers. The urea rapidly breaks down to carbamic acid and carbamylates the proteins, modifying their charge. Urea breakdown and subsequent protein carbamylation is the cause of charge trains on 2-D gels. A charge train is a series of spots on a 2-D gel that are of different pIs and the same size.

For more great tips visit www.expressionproteomics.com

Entry filed under: Miscellaneous Videos and Technotes, Proteomic Technotes. Tags: , , .

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