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The effects of high salt concentrations in the samples on molecular weight determination in sodium dodecyl sulfate polyacrylamide gel electrophoresis
Author(s) -
See Yew Phew,
Olley Peter M.,
Jackowski George
Publication year - 1985
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.1150060806
Subject(s) - chemistry , sodium dodecyl sulfate , chromatography , polyacrylamide gel electrophoresis , sodium , gel electrophoresis , electrophoresis , salt (chemistry) , acrylamide , ionic strength , phosphate , molecular mass , analytical chemistry (journal) , aqueous solution , biochemistry , enzyme , organic chemistry , monomer , polymer
For proper molecular weight determination in sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE), the protein should bind the same amount of SDS. High ionic strength had previously been shown to reduce the amount of SDS bound to proteins, suggesting that high salt concentrations in the samples may interfere with molecular weight determination. We examined the effects of NaCl from 0 to 1 M in the samples on molecular weight determination of proteins in SDS‐PAGE. In our conditions of electrophoresis, the retardation coefficients ( K R ), free mobilities ( Y o ) and R f values for proteins ( M r 20 000–66 000) are not affected by NaCl in the samples up to 0.8 M. Therefore molecular weight of these proteins can be determined by both the Ferguson plot and R f vs. log M r methods in the presence of NaCl in the samples up to 0.8 M and compared with standard protein samples in the absence of NaCl. The R f , K R and Y o values for phosphorylase ( M r 97 400) are affected by NaCl from 0.4 M or higher in the samples. Preliminary results show that increasing sodium phosphate, pH 7.0 from 0.01 M to 0.1 M in the sample affect molecular weight determinations. The effects of phosphate are greater in the Ferguson plot than in the R f vs. log M r plot procedure. In the latter method, the effects are dependent on the acrylamide gel concentrations and on the size of the proteins. We conclude that the molecular weight of proteins ( M r 20 000 to 97 000) can be determined in the presence of high salt concentration in the sample if the unknown and standard protein samples contain the same amount of salt and buffer ions.