Polyacrylamide lamination enables mass spectrometry compatible staining and in-gel digestion of proteins separated by agarose IEF.
Agarose IEF enables the separation of large proteins and protein complexes. A complication of agarose gels attached onto polyester support is the lack of sensitive protein staining methods compatible with protein analysis and identification protocols. In this study, agarose IEF gels were used to separate the proteins, followed by layering the agarose with polyacrylamide. The formed laminate gels were seamless and durable and they were readily detached from the polyester. The gels were amenable to MS compatible staining. The sensitivity obtained with the acidic silver staining method was 20-50 ng/band of myoglobin. Laminated agarose was a suitable matrix for in-gel digestion based generation of tryptic peptides for MALDI-MS.
Hellman
Jukka
J
University of Turku, Department of Biochemistry and Food Chemistry/Biotechnology, Turku, Finland. jukka.hellman@utu.fi
D000180:Acrylic Resins / Q000737:chemistry*; D000595:Amino Acid Sequence; D004587:Electrophoresis, Agar Gel / Q000379:methods*; D007525:Isoelectric Focusing / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods*; D008969:Molecular Sequence Data; D011506:Proteins / Q000032:analysis* / Q000235:genetics; D013194:Staining and Labeling / Q000379:methods*; D000099166:Polyacrylamides
D000180:Acrylic Resins; D011506:Proteins; D000099166:Polyacrylamides