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Performance of a novel sieving matrix of poly(vinyl alcohol)/acrylamide copolymer in electrophoretic separations of high molecular weight proteins from red cell membrane
Author(s) -
Matte Alessandro,
Sola Laura,
Chiari Marcella,
Tomelleri Carlo,
Consonni Roberto,
Turrini Franco,
Franceschi Lucia De
Publication year - 2014
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.201300529
Subject(s) - acrylamide , vinyl alcohol , polyacrylamide , copolymer , electrophoresis , chemistry , chromatography , matrix (chemical analysis) , polyacrylamide gel electrophoresis , membrane , chemical engineering , polymer chemistry , polymer , organic chemistry , biochemistry , engineering , enzyme
The analysis of high molecular weight (HMW) proteins from complex mixtures is still a challenge in proteomics. This work introduces a novel hydrogel obtained by the copolymerization of an allyl‐PVA derivative with acrylamide and bisacrylamide and applies this matrix to the electrophoretic separation of HMW proteins. By inducing gelation of polyacrylamide in the presence of variable amounts of allyl‐PVA, it is possible to control and vary the average gel porosity. This gel is easy to produce and handle and offers the advantage of being highly mechanically resistant and macroporous. The new matrix was tested in mono‐dimensional separations of complex protein mixtures extracted from red cell membranes with different detergents. The improved performance of this macroporous matrix allowed to identify new proteins by MS and immunoblot analysis using specific antibodies. In particular, the resolution of proteins ranging in size between 97 and 279 kDa was greatly improved here compared to standard polyacrylamide gels, suggesting that this matrix can be a useful tool in routine analysis of HMW proteins in cell biology.