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Preparative application of commercial automated gel electrophoresis apparatus to subcellular‐sized particles: Sequential isolations, fractions re‐run, sodium dodecyl sulfate‐polyacrylamide gel electrophoresis analysis, yield and purity
Author(s) -
Chen g,
Chrambach Andreas
Publication year - 1998
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.1150191810
Subject(s) - gel electrophoresis , sodium dodecyl sulfate , chromatography , electrophoresis , polyacrylamide gel electrophoresis , yield (engineering) , chemistry , two dimensional gel electrophoresis , gel electrophoresis of proteins , sodium , polyacrylamide , molecular weight size marker , color marker , biochemistry , proteomics , materials science , enzyme , gene , organic chemistry , metallurgy , polymer chemistry
The analytical and preparative potential of automated gel electrophoresis apparatus with intermittent fluorescence scanning of the migration path, the HPGE‐1000 apparatus (LabIntelligence, Belmont, CA) was further developed in application to subcellular‐sized particles. Resolution between two rat liver microsome components in agarose (MetaPhor) gel electrophoresis was found to increase with decreasing agarose concentration to 0.04%. It was less, even in an agarose solution at that low concentration, than that in laterally aggregated 4% polyacrylamide gel. The three components of the microsomal preparation were sequentially isolated from 0.6 and 0.8% agarose gel electropherograms. One fraction when reelectrophoresed was found to exhibit the original mobility and did not give rise to the other components. Yields of each component were near‐quantitative after one or two electroelution steps. Based on protein content, no impurities could be detected in two of the microsome fractions; the third fraction contained 2% of nonmicrosome impurity. Sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE) patterns of all three microsome fractions were indistinguishable from one another and from that of the unfractionated microsome preparation.