Triton-X 100 effect on protein concentration determination
John A. Berges
berges at BNLUX1.BNL.GOV
Fri Jul 7 08:49:55 EST 1995
>>>Effect of Triton X-100 on protein concentration determination.
>>>Has anyone in this net ever evaluated the effect of Triton-X 100 present in
>>protein sample when measuring protein
>>>concentration by the BioRad assay? If you did, please tell me what happened.
>>>Elida at parasit.lan.mcgill.ca
The effect of detergents on the Bradford (i.e. Biorad) protein assay is well
documented...it really messes things up. The absorbance of all your samples
will go sky-high. Biorad says up to about 0.1% Triton can be compensated
for by including the detergent in standards, but this leads to samples that
change very quickly in absorbance and precipitate within an hour or so.
Also, there are marked non-linearities in standard curves with Triton.
Biorad also has a detergent compatible assay, based on the Lowry procedure.
Bicinchoninic acid-based assays also work well with detergents; see kits by
Sigma, Pierce and others. These assays won't work in the presence of
reducing agents (e.g. DTT or mercaptoethanol).
Recently, Geno Technology has come out with a "dotMetric" assay that makes
some big promises in advertising (although the accompanying literature
qualifies many of these). It is supposed to overcome just about any
commoninterference. I haven't tried it yet...anyone else out ther have any
experience with it?
_/_/_/ _/ _/ _/ Dr. John A. Berges
_/ _/ _/ _/ _/ Oceanic and Atmospheric
_/ _/ _/_/ _/ _/ Sciences Divison
_/_/_/ _/ _/ _/ _/ Brookhaven National Lab
_/ _/ _/ _/ _/ _/ Upton, NY 11973
_/ _/ _/ _/_/ _/ Phone: 516-282-4077
_/_/_/_/ _/ _/ _/_/_/_/ Email: berges at BNLUX1.BNL.GOV
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