A new isoelectric focusing gel for two-dimensional electrophoresis constructed in microporous hollow fiber membranes.
We describe the preparation of IEF tube gels inside a nonwetting microporous plastic tubing. The gel in the tube need not be extruded after the first dimension separation. Instead, the porous structure of the tubes is made wettable, and the proteins are electrophoresed "through-the-wall" into the second dimension PAGE gel. Commercial ampholytes and reagents are suitable for the procedure. A useful p/ range of 4.5-9.5 can be obtained when p/ 3-10 ampholyte mixtures are used. Because of the high surface area of the porous material, precautions must be exercised to reduce oxygen inhibition during polymerization and dehydration of the gel during storage and use. A sheath device is described that satisfies these requirements. The plastic tubes can be disposed of by incineration and pose no biohazard.
Klein
Jon
J
Department of Medicine, the Core Proteomics Laboratory, University of Louisville School of Medicine, Louisville, Kentucky 40202, USA. jon.klein@louisville.edu
Harding
George
G
Klein
Elias
E
D000818:Animals; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D005347:Fibroblasts / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing / Q000379:methods*; D007526:Isoelectric Point; D008567:Membranes, Artificial; D011506:Proteins / Q000032:analysis* / Q000737:chemistry; D051381:Rats; D013997:Time Factors
D008567:Membranes, Artificial; D011506:Proteins