Premium
Two‐dimensional electrophoresis. Two‐dimensional gel electrophoresis of platelet polypeptides with immobilized pH gradients in capillary tubes
Author(s) -
Asakawa Junichi
Publication year - 1988
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.1150090916
Subject(s) - immobilized ph gradient , isoelectric focusing , chromatography , capillary electrophoresis , chemistry , electrophoresis , capillary action , analytical chemistry (journal) , materials science , biochemistry , composite material , enzyme
Two‐dimensional electrophoresis (2‐DE) with immobilized pH gradient (IPG) gels in capillary tubes was used in the first‐dimensional isoelectric focusing (IEF) for the separation of human platelet polypeptides. Two types of IPG tube gels, pH ranges 4–8 and 7–10, containing 8 M urea, 1% Nonidet P‐40 and 0.1% pH 3.5–10 Ampholine carrier ampholytes (CA) were prepared by a simple method not requiring special equipment. The addition of CA to both gel and sample solutions was essential in the tube gel IPG system. Proteins were visualized by a modification of Wray's silver‐staining technique. The degree of resolution and the number of spots observed on an IPG 2‐DE gel with pH 4–8 were comparable with those obtained with O'Farrell's high‐resolution 2‐DE. Approximately 200 basic polypeptides, which are difficult to separate by conventional CA‐based IEF 2‐DE or the non‐equilibrium pH gradient system, were well resolved by 2‐DE with a pH 7–10 IPG tube gel in the first‐dimension. The gel patterns with either pH gradient 4–8 or 7–10 were highly reproducible among gels prepared and run simultaneously. These results demonstrated the potential and usefulness of the 2‐DE system with IPG gels in capillary tubes.