Application of plasma-polymerized films for isoelectric focusing of proteins in a capillary electrophoresis chip.

The first use of plasma polymerization technique to modify the surface of a glass chip for capillary isoelectric focusing (cIEF) of different proteins is reported. The electrophoresis separation channel was machined in Tempax glass chips with length 70 mm, 300 microm width and 100 microm depth. Acetonitrile and hexamethyldisiloxane monomers were used for plasma polymerization. In each case 100 nm plasma polymer films were coated onto the chip surface to reduce protein wall adsorption and minimize the electroosmotic flow. Applied voltages of 1000 V, 2000 V and 3000 V were used to separate mixtures of cytochrome c (pI 9.6), hemoglobin (pI 7.0) and phycocyanin (pI 4.65). Reproducible isoelectric focusing of each pI marker protein was observed in different coated capillaries at increasing concentration 2.22-5 microg microL(-1). Modification of the glass capillary with hydrophobic HMDS plasma polymerized films enabled rapid cIEF within 3 min. The separation efficiency of cytochrome c and phycocyanin in both acrylamide and HMDS coated capillaries corresponded to a plate number of 19600 which compares favourably with capillary electrophoresis of neurotransmitters with amperometric detection.

[1]  D. J. Harrison,et al.  Micromachining a Miniaturized Capillary Electrophoresis-Based Chemical Analysis System on a Chip , 1993, Science.

[2]  Andreas Manz,et al.  High-Speed Separation of Antisense Oligonucleotides on a Micromachined Capillary Electrophoresis Device , 1994 .

[3]  T. Wehr,et al.  Performance of Uncoated and Coated Capillaries in Free Zone Electrophoresis and Isoelectric Focusing of Proteins , 1991 .

[4]  S. Hjertén,et al.  Theoretical and experimental study of high-performance electrophoretic mobilization of isoelectrically focused protein zones. , 1987, Journal of chromatography.

[5]  D. J. Harrison,et al.  Microchip-based capillary electrophoresis of human serum proteins. , 1997, Journal of chromatography. A.

[6]  S. Hjertén,et al.  Separation of the human transferrin isoforms by carrier-free high-performance zone electrophoresis and isoelectric focusing. , 1989, Journal of chromatography.

[7]  A. Chen,et al.  Carrier-free zone electrophoresis, displacement electrophoresis and isoelectric focusing in a high-performance electrophoresis apparatus. , 1987, Journal of chromatography.

[8]  J Wang,et al.  Integrated electrophoresis chips/amperometric detection with sputtered gold working electrodes. , 1999, Analytical chemistry.

[9]  I. Karube,et al.  Mass transport behavior of electrochemical species through plasma-polymerized thin film on platinum electrode , 2000 .

[10]  Stellan Hjertén,et al.  Adaptation of the equipment for high-performance electrophoresis to isoelectric focusing , 1985 .

[11]  R. McCormick.  Capillary zone electrophoretic separation of peptides and proteins using low pH buffers in modified silica capillaries. , 1988, Analytical chemistry.

[12]  Stellan Hjertén,et al.  High-performance electrophoresis : Elimination of electroendosmosis and solute adsorption , 1985 .

[13]  A. Manz,et al.  Glass chips for high-speed capillary electrophoresis separations with submicrometer plate heights , 1993 .

[14]  S. Hjertén,et al.  Fast and high resolution analysis of human serum transferrin by high performance isoelectric focusing in capillaries , 1989, Electrophoresis.

[15]  I. Karube,et al.  Application of polymer-embedded proteins to fabrication of DNA array. , 2000, Biotechnology and bioengineering.

[16]  M. Chiari,et al.  Characterization of poly(dimethylacrylamide) and the combination of poly(vinyl alcohol) and cetyltrimethylammonium bromide as dynamic electroosmotic flow suppression agents in capillary electrophoresis , 1998 .

[17]  O Hofmann,et al.  Adaptation of capillary isoelectric focusing to microchannels on a glass chip. , 1999, Analytical chemistry.

[18]  J. Michael Ramsey,et al.  Precolumn Reactions with Electrophoretic Analysis Integrated on a Microchip , 1994 .

[19]  D. J. Harrison,et al.  Planar chips technology for miniaturization and integration of separation techniques into monitoring systems. Capillary electrophoresis on a chip , 1992 .

[20]  J. Michael Ramsey,et al.  Microchip Capillary Electrophoresis with an Integrated Postcolumn Reactor , 1994 .

[21]  S. Jacobson,et al.  Integrated microdevice for DNA restriction fragment analysis. , 1996, Analytical chemistry.

[22]  H. Muguruma.  Plasma-polymerized films for biosensors , 1999 .

[23]  Richard A. Mathies,et al.  Capillary array electrophoresis: an approach to high-speed, high-throughput DNA sequencing , 1992, Nature.

[24]  Andreas Manz,et al.  Planar glass chips for capillary electrophoresis: repetitive sample injection, quantitation, and separation efficiency , 1993 .

[25]  J. Pawliszyn,et al.  Dynamics of capillary isoelectric focusing in the absence of fluid flow: high-resolution computer simulation and experimental validation with whole column optical imaging. , 2000, Analytical chemistry.

[26]  D. J. Harrison,et al.  Capillary electrophoresis and sample injection systems integrated on a planar glass chip , 1992 .

[27]  Janusz Pawliszyn,et al.  Demonstration of isoelectric focusing on an etched quartz chip with UV absorption imaging detection , 1999 .

[28]  J. Adamovics,et al.  SEPARATION OF THE ISOFORMS OF A MONOCLONAL ANTIBODY BY GEL ISOELECTRIC FOCUSING, HIGH PERFORMANCE LIQUID CHROMATOGRAPHY AND CAPILLARY ISOELECTRIC FOCUSING , 1992 .

[29]  M. Novotny,et al.  Electrophoretic separations of proteins in capillaries with hydrolytically stable surface structures. , 1990, Analytical chemistry.

[30]  N. Guzman.  Capillary electrophoresis technology , 1993 .

[31]  I Karube,et al.  A novel method of immobilizing antibodies on a quartz crystal microbalance using plasma-polymerized films for immunosensors. , 1996, Analytical chemistry.