Contributors: J. Ou, T. Glawdel, R. Samy, C. L. Ren, Z. Liu, T. Huang, J. Pawliszyn
Isoelectric focusing (IEF) as one of the high-resolution capillary electrophoresis (CE) techniques has been widely applied for the separation of zwitterionic biomolecules, such as proteins and peptides, based on the differences of their isoelectric points (pIs). Whole-column imaging detection (WCID) developed by Pawliszyn et al is an ideal detection method for IEF because no mobilization is required. This research project aims to develop novel microfluidic chips for IEF and miniaturized WCID techniques. The intended main objectives of this project includes three phases.
1) The first phase involves design and fabrication of a microfluidic chip with inexpensive material, polydimethylsiloxane (PDMS). The new PDMS microfluidic chip-based cartridge can be an available alternative to the standard fused-silica capillary-based cartridge used for IEF-WCID analysis in a commercial instrument, iCE280 Analyzer (Convergent Bioscience, Toronto).
Top view (a), side view (b) and photograph (c) of a 100 µm x 100 µm x 5 cm-channel PDMS microchip.
2) The second involves development of innovative WCID technologies to make the system more sensitive, compact and inexpensive. The microfluidic chip described above will be modified to allow for the operation of ultraviolet (UV) absorption and laser-induced fluorescence (LIF) WCID on a single run.
Capillary isoelectric focusing (CIEF) profile of human hemoglobin control AFSC on (a) a microchip and (b) a capillary cartridge. Separation channel: (a) 100 µm x 100 µm x 5 cm; (b) 100 µm ID x 5 cm. Sample: 0.1 mg/mL human hemoglobin control containing 2% Pharmalytes (pH 3-10) and 1% PVP.
3) The third phase will focus on development of microfluidic chip-based two-dimensional CE separations (2D-CE) and integration of chip with mass spectrometry (MS).
Bibliography
J. Ou, T. Glawdel, R. Samy, S. Wang, Z. Liu, C. L. Ren, J. Pawliszyn “Integration of dialysis membranes into a poly(dimethylsiloxane) microfluidic chip for isoelectric focusing of proteins using whole-channel imaging detection (PDF)”, Anal Chem, 80 (2008) 7401-7407.
J. Ou, T. Glawdel, C. L. Ren, J. Pawliszyn, “Fabrication of a Hybrid PDMS/SU-8/quartz Microfluidic Chip for Enhancing UV Absorption Whole-channel Imaging Detection Sensitivivity and Application for Isoelectric Focusing of Proteins (PDF)”, Lab Chip, accepted, DOI:10.1039/B821438G.