Free-Flow Zone Electrophoresis of Peptides and Proteins in PDMS Microchip for Narrow pI Range Sample Prefractionation Coupled with Mass Spectrometry

• Main Authors: Chan, Michael (Author), Song, Yong-Ak (Contributor), Celio, Chris (Contributor), Tannenbaum, Steven Robert (Contributor), Wishnok, John S. (Contributor), Han, Jongyoon (Contributor)
• Other Authors: Massachusetts Institute of Technology. Department of Biological Engineering (Contributor), Massachusetts Institute of Technology. Department of Chemistry (Contributor), Massachusetts Institute of Technology. Department of Electrical Engineering and Computer Science (Contributor), Massachusetts Institute of Technology. Research Laboratory of Electronics (Contributor), Tannenbaum, Steven R. (Contributor)
• Format: Article
• Language: English
• Published: American Chemical Society, 2011-11-02T19:47:12Z.
• Subjects: Article
• Online Access: Get fulltext

In this paper, we are evaluating the strategy of sorting peptides/proteins based on the charge to mass without resorting to ampholytes and/or isoelectric focusing, using a single- and two-step free-flow zone electrophoresis. We developed a simple fabrication method to create a salt bridge for free-flow zone electrophoresis in PDMS chips by surface printing a hydrophobic layer on a glass substrate. Since the surface-printed hydrophobic layer prevents plasma bonding between the PDMS chip and the substrate, an electrical junction gap can be created for free-flow zone electrophoresis. With this device, we demonstrated a separation of positive and negative peptides and proteins at a given pH in standard buffer systems and validated the sorting result with LC/MS. Furthermore, we coupled two sorting steps via off-chip titration and isolated peptides within specific pI ranges from sample mixtures, where the pI range was simply set by the pH values of the buffer solutions. This free-flow zone electrophoresis sorting device, with its simplicity of fabrication, and a sorting resolution of 0.5 pH unit, can potentially be a high-throughput sample fractionation tool for targeted proteomics using LC/MS.
Korea Institute of Science and Technology. Intelligent Microsystems Center
Massachusetts Institute of Technology. Center for Environmental Health Sciences
National Institute of Environmental Health Sciences (Grant No. P30-ES002109)
United States. National Institutes of Health (grant R21 EB008177)