Separating the many components of a proteomic sample is very challenging due to the complexity of the mixture. Protein expression at the cellular level is challenging and the dynamic range of some proteins can range from 106 to 109 per cell. Therefore, there is still a need for separations with higher peak capacity, better automation and faster run times in order to improve proteomic analyses as well as throughput. A single analysis permitting the identification of the entire proteome is typically difficult because of the complexity and the limited resolution of 2-D electrophoresis and the limited dynamic range and limit-of-detection of mass spectrometry. The fluidic system will consist of 4 modules (1) Cell biotinylation and lysis made from polycarbonate (PC) due to its high glass transition temperature and compatibility with the PC micro-membrane which has a specific pore size that aids in allowing only the biotinylated cells to permeate; (2) solid-phase affinity extraction module made from poly(methyl methacrylate) (PMMA) due to its ease of UV-surface modification, biocompatibility and high surface density of functional groups; (3) 2-D µCE module also made from PMMA because it has been demonstrated to generate high plate numbers for electrophoretic separations of proteins; and (4) solid-phase bioreactor, also made of PMMA, for proteolytic digestion of protein components sorted via 2-D µCE. Each module, as well as the fluidic motherboard, will be made from thermoplastics via a metal molding tool used to replicate the prerequisite parts by hot embossing. Chips will be molded using metal molding tools fabricated via high precision micromilling (HPMM) or UV-LiGA. The fluidic network will be enclosed using thermal fusion bonding. This work is supported by the National Science Foundation’s Graduate Research Fellowship.
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