Protein concentration by precipitation with pyrogallol red prior to electrophoresis
作者:
Thomas Marshall,
Nicola J. Abbott,
Peter Fox,
Katherine M. Williams,
期刊:
ELECTROPHORESIS
(WILEY Available online 1995)
卷期:
Volume 16,
issue 1
页码: 28-31
ISSN:0173-0835
年代: 1995
DOI:10.1002/elps.1150160107
出版商: Wiley Subscription Services, Inc., A Wiley Company
关键词: Protein concentration;Pyrogallol red;Urinary proteins
数据来源: WILEY
摘要:
AbstractThe pyrogallol red protein assay (Clinical Chemistry1986,32, 1551–1554) is based upon formation of a blue protein‐dye complex in the presence of molybdate under acidic conditions. However, centrifugation of the assay mixture results in loss of color yield and precipitation of the protein‐dye complex which can be recovered and resolubilized to achieve protein concentration prior to sodium dodecyl sulfate‐polyacrylamide gel electrophoresis. The method has been evaluated relative to trichloroacetic acid (TCA) precipitation for recovery and electrophoresis of commercial protein and peptide molecular weight markers. Precipitation with pyrogallol red‐molybdate (PRM) gives better and more uniform recovery of both proteins and peptides as compared to TCA. The lower limit of PRM precipitation is similar to TCA and corresponds to 1 μg protein per mL assay mixture. This is equivalent to 100 μL of 10 μg/mL protein using the standard protein assay or 1 μg/mL protein using a modified assay incorporating a fivefold concentrate of the dye reagent. Application of the method is demonstrated by concentration of urinary proteins. The method is simple and economic and useful for conserving trace amounts of precious sample as it allows recovery of protein for electrophoresis following
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