Biomedical Microdevices

, Volume 9, Issue 4, pp 533–543 | Cite as

Automatic microfluidic platform for cell separation and nucleus collection

  • Chien-Hsuan Tai
  • Suz-Kai Hsiung
  • Chih-Yuan Chen
  • Mei-Lin Tsai
  • Gwo-Bin Lee
Article

Abstract

This study reports a new biochip capable of cell separation and nucleus collection utilizing dielectrophoresis (DEP) forces in a microfluidic system comprising of micropumps and microvalves, operating in an automatic format. DEP forces operated at a low voltage (15 Vp–p) and at a specific frequency (16 MHz) can be used to separate cells in a continuous flow, which can be subsequently collected. In order to transport the cell samples continuously, a serpentine-shape (S-shape) pneumatic micropump device was constructed onto the chip device to drive the samples flow through the microchannel, which was activated by the pressurized air injection. The mixed cell samples were first injected into an inlet reservoir and driven through the DEP electrodes to separate specific samples. Finally, separated cell samples were collected individually in two outlet reservoirs controlled by microvalves. With the same operation principle, the nucleus of the specific cells can be collected after the cell lysis procedure. The pumping rate of the micropump was measured to be 39.8 μl/min at a pressure of 25 psi and a driving frequency of 28 Hz. For the cell separation process, the initial flow rate was 3 μl/min provided by the micropump. A throughput of 240 cells/min can be obtained by using the developed device. The DEP electrode array, microchannels, micropumps and microvalves are integrated on a microfluidic chip using micro-electro-mechanical-systems (MEMS) technology to perform several crucial procedures including cell transportation, separation and collection. The dimensions of the integrated chip device were measured to be 6 × 7 cm. By integrating an S-shape pump and pneumatic microvalves, different cells are automatically transported in the microchannel, separated by the DEP forces, and finally sorted to specific chambers. Experimental data show that viable and non-viable cells (human lung cancer cell, A549-luc-C8) can be successfully separated and collected using the developed microfluidic platform. The separation accuracy, depending on the DEP operating mode used, of the viable and non-viable cells are measured to be 84 and 81%, respectively. In addition, after cell lysis, the nucleus can be also collected using a similar scheme. The developed automatic microfluidic platform is useful for extracting nuclear proteins from living cells. The extracted nuclear proteins are ready for nuclear binding assays or the study of nuclear proteins.

Keywords

Dielectrophoresis force Micropump Microvalve Cell separation Cell collection MEMS Microfluidics 

Nomenclature

AC

Alternating current

Bio-MEMS

Bio-micro-electro-mechanical-systems

DEP

Dielectrophoresis

DMEM

Dulvecco’s modified eagle medium

DNA

Deoxyribonucleic acid

EMV

Electromagnetic valve

ER

Estrogen nuclear receptor

LIF

Laser induced fluorescence

LOC

Lab-on-a-chip

MEMS

Micro-electro-mechanical-systems

PDMS

Polydimethylsiloxane

PI

Propidium iodide

PR

Photoresist

SEM

Scanning electron microscope

S-shape

Serpentine-shape

Vp–p

Peak-to-peak Voltage

Notes

Acknowledgements

The authors gratefully acknowledge the financial support provided to this study by the National Science Council in Taiwan and by the MOE Program for Promoting Academic Excellence of Universities (EX-91-E-FA09-5-4). Also, the access provided to major fabrication equipment at the Center for Micro/Nano Technology Research, National Cheng Kung University is greatly appreciated.

References

  1. J.R. Anderson, D.T. Chiu, R.J. Jackman, O. Cherniavskaya, J.C. McDonald, H. Wu, S.H. Whitesides, G.M. Whitesides, Anal. Chem. 72, 3158 (2000)CrossRefGoogle Scholar
  2. J. Auerswald, H.F. Knapp, Microelectron. Eng. 67–68, 879 (2003)CrossRefGoogle Scholar
  3. P.A. Auroux, D.R. Reyes, D. Iossifidis, A. Manz, Anal. Chem. 74, 2637 (2002)CrossRefGoogle Scholar
  4. A.S. Bahaj, A.G. Bailey, in Proceedings of the Industry Applications Society (IEEE) Annual Meeting, Cleveland, OH, 1979, p. 154Google Scholar
  5. F.F. Becker, X.B. Wang, Y. Huang, R. Pethig, J. Vykoukal, P.R.C. Gascoyne, Proc. Natl. Acad. Sci. U. S. A. 92, 860 (1995)CrossRefGoogle Scholar
  6. M. Brown, C. Wittwer, Clin. Chem. 46, 1221 (2000)Google Scholar
  7. N.H. Chiem, D.J. Harrison, Clin. Chem. 44, 591 (1998)Google Scholar
  8. Y.J. Chuang, M.L. Tsai, S.H. Chen, Electrophoresis 27, 4158 (2006)CrossRefGoogle Scholar
  9. I. Doh, Y.H. Cho, Sens. Actuators A 121, 59 (2005)CrossRefGoogle Scholar
  10. D.C. Duffy, J.C. McDonald, O.J.A. Schueller, G.M. Whitesides, Anal. Chem. 70, 4974 (1998)CrossRefGoogle Scholar
  11. B.S. Edwards, T. Oprea, E.R. Prossnitz, L.A. Sklar, Curr. Opin. Chem. Biol. 8, 392 (2004)CrossRefGoogle Scholar
  12. L.M. Fu, G.B. Lee, Y.H. Lin, R.J. Yang, IEEE/ASME Trans. Mechatron. 9, 377 (2004)CrossRefGoogle Scholar
  13. P.R.C. Gascoyne, X.B. Wang, Y. Huang, F.F. Becker, IEEE Trans. Ind. Appl. 33, 670 (1997)CrossRefGoogle Scholar
  14. C.F. Gonzalez, V.T. Remcho, J. Chromatogr. A 1079, 59 (2005)CrossRefGoogle Scholar
  15. Y. Huang, R. Holzel, R. Pethig, X.B. Wang, Phys. Med. Biol. 37, 1499 (1992)CrossRefGoogle Scholar
  16. Y. Huang, K.L. Ewalt, M. Tirado, R. Haigis, A. Forster, D. Ackley, M.J. Heller, J.P. O’Connel, M. Krihak, Anal. Chem. 73, 1549 (2001)CrossRefGoogle Scholar
  17. K.K. Jaln, Trends Biotechnol. 18, 278 (2000)CrossRefGoogle Scholar
  18. T.B. Jones, Electromechanics of Particles (Cambridge University Press, New York, 1995)Google Scholar
  19. T.B. Jones, IEEE Proc. Nanobiotechnol. 39, 150 (2003)Google Scholar
  20. D.J. Laser, J.G. Santiago, J. Micromechanics Microengineering 14, 35 (2004)CrossRefGoogle Scholar
  21. G.B. Lee, C.H. Lin, S.C. Chang, J. Micromechanics Microengineering 15, 447 (2005)CrossRefGoogle Scholar
  22. H. Li, R. Bashir, Sens. Actuators B 86, 215 (2002)CrossRefGoogle Scholar
  23. C.H. Lin, G.B. Lee, B.W. Chang, G.L. Chang, J. Micromechanics Microengineering 12, 590 (2002)CrossRefGoogle Scholar
  24. A. Manz, D.J. Harrison, J.C. Verpoorte, H. Ludi, H.M. Widmer, in Technical Digest IEEE Transducers ’91, San Francisco, 1990, p. 939Google Scholar
  25. G.H. Markx, M.S. Talary, R. Pethig, J. Biotechnol. 32, 29 (1994)CrossRefGoogle Scholar
  26. H.A. Pohl, J. Appl. Phys. 22, 869 (1951)CrossRefGoogle Scholar
  27. H.A. Pohl, J. Appl. Phys. 29, 1182 (1958)CrossRefGoogle Scholar
  28. H.A. Pohl, Dielectrophoresis (Cambridge University Press, Cambridge, UK, 1978)Google Scholar
  29. R. Raiteri, M. Grattarola, R. Berger, Materials Today 5, 22 (2002)CrossRefGoogle Scholar
  30. D.R. Reyes, D. Iossifidis, P.A. Auroux, A. Manz, Anal. Chem. 74, 2623 (2002)CrossRefGoogle Scholar
  31. K. Sato, A. Hibara, M. Tokeshi, H. Hisamoto, T. Kitamori, Adv. Drug Deliv. Rev. 55, 379 (2003)CrossRefGoogle Scholar
  32. M. Rieseberg, C. Kasper, K.F. Reardon, T. Scheper, Appl. Microbiol. Biotechnol. 56, 350 (2001)CrossRefGoogle Scholar
  33. A. Ring, M. Dowsett, Endocr.-Relat. Cancer 11, 643 (2004)CrossRefGoogle Scholar
  34. J. Rousselet, G.H. Markx, R. Pethig, Colloids Surf. A 140, 209 (1998)CrossRefGoogle Scholar
  35. T. Schnelle, T. Müller, G. Gradl, S.G. Shirley, G. Fuhr, Electrophoresis 21, 66 (2000)CrossRefGoogle Scholar
  36. P. Skehan, R. Storeng, D. Scudiero, A. Monks, J. McMahon, D. Vistica, J.T. Warren, H. Bokesch, S. Kenney, M.R. Boyd, J. Natl. Cancer Inst. 82, 1107 (1990)CrossRefGoogle Scholar
  37. P.L. Tazzari, A. Bontadini, F. Fruet, C. Tassi, F. Ricci, S. Manfroi, R. Conte, Vox Sang. 85, 109 (2003)CrossRefGoogle Scholar
  38. M.A. Unger, H.P. Chou, T. Thorsen, A. Scherer, S.R. Quake, Science 288, 113 (2000)CrossRefGoogle Scholar
  39. C.H. Wang, G.B. Lee, Biosens. Bioelectron. 21, 419 (2005)CrossRefGoogle Scholar
  40. C.H. Wang, G.B. Lee, J. Micromechanics Microengineering 16, 341 (2006)CrossRefGoogle Scholar
  41. X.B. Wang, Y. Huang, P.R.C. Gascoyne, F.F. Becker, R. Holzel, R. Pethig, Biochim. Biophys. Acta 1193, 330 (1994)CrossRefGoogle Scholar
  42. R.J. Widrow, C.D. Laird, Cytometry 39, 126 (2000)CrossRefGoogle Scholar
  43. R.J. Widrow, P.S. Rabinovitch, K. Cho, C.D. Laird, Cytometry 27, 250 (1997)CrossRefGoogle Scholar
  44. S.Y. Yang, S.K. Hsiung, Y.C. Hung, C.M. Chang, T.L. Liao, G.B. Lee, Meas. Sci. Technol. 17, 2001 (2006)CrossRefGoogle Scholar
  45. R. Zengerle, J. Ulrich, S. Kluge, M. Richter, A. Richter, Sens. Actuators A 50, 81 (1995)CrossRefGoogle Scholar
  46. B. Ziaie, A. Baldi, M. Lei, Y. Gu, R.A. Siegel, Adv. Drug Deliv. Rev. 56, 145 (2004)CrossRefGoogle Scholar
  47. H. Zou, S. Mellon, R.R. Syms, K.E. Tanner, Biomed. Microdevices 8, 353 (2006)CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2007

Authors and Affiliations

  • Chien-Hsuan Tai
    • 1
  • Suz-Kai Hsiung
    • 1
  • Chih-Yuan Chen
    • 2
  • Mei-Lin Tsai
    • 2
  • Gwo-Bin Lee
    • 1
  1. 1.Department of Engineering ScienceNational Cheng Kung UniversityTainanTaiwan
  2. 2.Department of PhysiologyNational Cheng Kung UniversityTainanTaiwan

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