Patent ID: 8728290

Claim:
A method for rapid two-dimensional (2-1)) separation of proteins and related polypeptide molecules, comprising the steps of: providing a microfluidic device comprising a first microchannel and a plurality of branching side microchannels each intersecting and in fluid communication with the first microchannel, wherein the first microchannel and each of the branching side microchannels comprise one or more interior surfaces comprising an acrylate-terminated coating covalently bonded thereto, and a length and width, wherein all of the branching side microchannels are in fluid communication; forming an IEF stage within the first microchannel, wherein the step of forming the IEF stage comprises the steps of: introducing an acrylamide/bisacrylamide solution into a length of the first microchannel and forming a pH gradient therein; and polymerizing the acrylamide/bisacrylamide solution across a partial width or depth of a length of the one branching side microchannel, wherein the porous gel is covalently bonded to the acrylate-terminated coating within a length of the first microchannel, and wherein the remaining unpolymerized acrylamide/bisacrylamide solution is removed; forming a PAGE stage within at least some of the plurality of branching side microchannels; introducing a sample analyte comprising a plurality of proteins and related polypeptide molecules into a loading microchannel communicating directly with the first microchannel; applying a first electromotive force across a first and a second end of the IEF stage thereby causing the proteins and related polypeptide molecules within the sample analyte to separate into a plurality of bands located at isoelectric points (pi) along the length of the IEF stage; and applying a second electromotive force across first and second ends of each of the branching side microchannels comprising the PAGE stage thereby causing the plurality of bands in the IEF stage to further separate into one or more refined bands along the length of each of the one or more branching side microchannels comprising the PAGE stage.