Gel gradient electrophoresis, isoelectric focusing and two-dimensional techniques in horizontal, ultrathin polycrylamide layers

Abstract

A ultrathin layer, horizontal polycrylamide gel system for electrophoresis, isoelectric focusing and two-dimensional techniques is described. Gel slabs 240 μm thin for unidimensional, or 360 μm thin for two-dimensional runs are cast on cellphone foils as support. The sample is loaded in pockets pre-cast in the gel (2–3 μl size) or in trenches for two-dimensional experiments. The second dimension is routinely performed in concave exponential gel gradients, spanning an acrylamide concentration from 4% to 22.5%. The sensitivity with the common Coomassie Blue stain is very high, well below 0.1 μg protein/band. Zymogram detections can be developed within a few minutes, thus retaining the band sharpness of the focused zones or of the bands separated in pore gradient electrophoresis. Sample handling, stainingand destaining and gel drying and storage are greatly simplified and performed in a fraction of the time needed for conventional, thick gels in the 1–2 mm thickness range.