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A new method for the preperative and analytical electrophoresis of cells


In this paper, a new method is described for the horizontal electrophoresis of cells on a density cushion under near-isopycnic conditions. When cell sedimentation is minimized, the electrophoresis of red blood cells (RBC) used as model cells within an anti-convective porous matrix (with pores over 300 μm in diameter) was capable of separating a mixture of human and chicken RBC according to their electrophoretic mobilities. Samples taken from the separated RBC bands show over 90% purity for each species. The simultaneous electrophoresis of several RBC samples carried out under identical conditions permitted the use of comparative data based on the electrophoretic mobility of cells which differ in their surface properties. We believe that this relatively simple system, in which cell sedimentation and convection are minimized, has the potential to be modified and adapted for the separation of other cell types/organelles.



direct current electric field


molecular weight


phosphate buffered saline


polyethylene glycol


red blood cells


  1. 1.

    Abramson, H.A., Moyer, L.S. and Goris, M.H. Electrophoresis of proteins and the chemistry of cell surfaces, Reinhold NY 1942, 1-307.

  2. 2.

    Blanco, S., Clifton, M.J., Joly, J.L. and Peltre, G. Protein separation by electrophoresis in nonsieving amphoteric medium. Electrophoresis 17 (1996) 1126–1133.

    PubMed  CAS  Article  Google Scholar 

  3. 3.

    Chiari, M. and Righetti, P.G. New types of separation matrices for electrophoresis. Electrophoresis 16 (1995) 1815–1829.

    PubMed  CAS  Article  Google Scholar 

  4. 4.

    Roman, M.C. and Brown, P.R. Free-flow electrophoresis as a preparative separation technique. Anal. Chem. 66 (1994) 86–94.

    Google Scholar 

  5. 5.

    Wang, Y., Hancock, W.S., Weber, G., Eckerskorn, C. and Palmer-Toy, D. Free-flow electrophoresis coupled with liquid chromatography/mass spectrometry for a proteomic study of the human cell line (K562/CR3). J. Chromatogr. A 1053 (2004) 269–278.

    PubMed  CAS  Article  Google Scholar 

  6. 6.

    Zakharov, S.F., Chang, H.T. and Chrambach, A. Reproducibility of mobility in gel electrophoresis. Electrophoresis 17 (1996) 84–90.

    PubMed  CAS  Article  Google Scholar 

  7. 7

    Ambrose, E.J. Cell Electrophoresis. J&A Churchill Ltd., London 1965.

    Google Scholar 

  8. 8.

    Chaubal, K.A. Cell electrophoretic mobility as an aid to study biological systems, in: Cell Electrophoresis (Schütt, W., Klinkmann, H., Ed.), Walter de Gruyter, Berlin (NY) 1985, 515–526.

    Google Scholar 

  9. 9.

    Fürész, J., Pál, K., Budavári, I. and Lapis, K. The physicochemical properties of tumor cells with different metastatic potential. Neoplasma 32 (1985) 689–693.

    PubMed  Google Scholar 

  10. 10.

    Korohoda, W. Electrophoretic studies on plant cells III. Electrophoretic mobilities of cell-forms of Myxomycetae Physarum nudum Macbride. Folia Biologica 11 (1963) 465–472.

    Google Scholar 

  11. 11.

    Mehrishi, J.N. and Bauer, J. Electrophoresis of cells and the biological relevance of surface charge. Electrophoresis 23 (2002) 1984–1994.

    PubMed  CAS  Article  Google Scholar 

  12. 12.

    Mori, T. and Shimizu, M. The changes of lymphocyte electrophoretic mobility in cancer patient. in: Cell Electrophoresis (Schütt, W., Klinkmann, H., Ed.), Walter de Gruyter, Berlin (NY), 1985, 355–366.

    Google Scholar 

  13. 13.

    Preece, A.W. and Sablovic, D. in: Cell electrophoresis: clinical application and methodology. North-Holland Publishing company, Amsterdam 1979.

    Google Scholar 

  14. 14.

    Abercrombie, M. and Ambrose, E.J. The surface properties of cancer cells: a review. Cancer Res. 22 (1962) 332–245.

    Google Scholar 

  15. 15.

    Jovtchev, S., Djenev, I., Stoeff, S. and Stoylov, S. Role of electrical and mechanical properties of red blood cells for their aggregation. Colloids and Surfaces A: Physicochem. Engineer. Asp. 164 (2000) 95–104.

    CAS  Article  Google Scholar 

  16. 16.

    Vransky, V.K. Die zellelektrophorese. in: Fortschritte der experimentellen und theoretischen Biophysik Band 18. (Beier, W., Ed.) Leipzig 1974.

  17. 17.

    Masui, M., Takata, H. and Kominami, T. Cell adhesion and negative cell surface charges in embryonic cells of the starfish Asterina pectinifera. Electrophoresis 23 (2002) 2087–2095.

    PubMed  CAS  Article  Google Scholar 

  18. 18.

    Platsoucas, C.D., Good, R.A. and Gupta, S. Separation of human T lymphocyte subpopulations (Tμ, Tγ) by density gradient electrophoresis. Proc. Natl. Acad. Sci. USA 76 (1979) 1972–1976.

    PubMed  CAS  Article  Google Scholar 

  19. 19.

    Rychly, J., Anders, O., Eggers, G. and Schulz, M. Electrophoretic mobility distribution of cells in leukaemia. in: Cell Electrophoresis (Schütt, W., Klinkmann, H., Ed.), Walter de Gruyter, Berlin (NY) 1985, 477–483.

    Google Scholar 

  20. 20.

    Heidrich, H.G. and Hannig, K. Separation of cell population by free-flow electrophoresis. Methods Enzymol. 171 (1989) 513–531.

    PubMed  CAS  Article  Google Scholar 

  21. 21.

    Rutishauser, U.S. and Edelman, G.M. Fractionation and Manipulation of Cells with Chemically Modified Fibers and Surfaces. in: Methods of Cell Separation, vol. 1. (Catsimpoolas, N., Ed.) Plenum Press NY 1977, 193–228.

    Google Scholar 

  22. 22.

    Sengeløv, H. and Borregaard, N. Free-flow electrophoresis in subcellular fractionation of human neutrophils. J. Immunol. Methods 232 (1999) 145–152.

    PubMed  Article  Google Scholar 

  23. 23.

    Neu, B., Armstrong, J.K., Fisher, T.C. and Meiselman, H.J. Surface characterization of poly(ethylene glycol) coated human red blood cells by particle electrophoresis. Biorheology 40 (2003) 477–487.

    PubMed  CAS  Google Scholar 

  24. 24.

    Seaman, G.V.F. and Cook, G.M.W. Modification of the electrophoretic behavior of the erythrocyte by chemical and enzymatic methods. in: Cell Electrophoresis (Ambrose, E.J., Ed.). J&A Churchill Ltd., London 1965, 48–65.

    Google Scholar 

  25. 25.

    Wilson, W.W., Wade, M.M., Holman, S.C. and Champlin, F.R. Status of methods for assessing bacterial cell surface charge properties based on zeta potential measurements. J. Microbiol. Methods 43 (2001) 153–164.

    PubMed  CAS  Article  Google Scholar 

  26. 26.

    Catsimpoolas, N. and Griffith, A.L. Transient electrophoresis and sedimentation analyses of cells in density gradients. In: Methods of Cell Separation, vol. 2. (Catsimpoolas, N., Ed.) Plenum Press NY 1979, 1–63.

    Google Scholar 

  27. 27.

    Pertoft, H. and Lauren, T.C. Isopycinc separation of cells and cell organelles by centrifugation in modified colloidal silica gradients. in: Methods of Cell Separation, vol. 1. (Catsimpoolas, N., Ed.) Plenum Press NY 1977, 25–65.

    Google Scholar 

  28. 28.

    PretlowII, T.G. and Pretlow, T.P. Separation of viable cells by velocity sedimentation in an isokinetic gradient of ficoll in tissue culture medium. in: Methods of Cell Separation, vol. 1. (Catsimpoolas, N., Ed.) Plenum Press NY 1977, 171–191.

    Google Scholar 

  29. 29.

    Akiba, T., Nishi, A., Takaoki, M., Matsumiya, H., Tomita, F., Usami, R. and Nagaoka, S. Separation of bacterial cells by free flow electrophoresis under microgravity: a result of the spacelab — Japan project on space shuttle flight sts — 47. Acta Astron. 36 (1995) 177–181.

    CAS  Article  Google Scholar 

  30. 30.

    Zeiller, K., Löser, R., Pascher, G. and Hannig, K. Free-flow electrophoresis II: Analysis of the method with respect to preparative cell separation. Hoppe-Seyler’s Z Physiol. Chem. 356 (1975) 1225–1244.

    PubMed  CAS  Google Scholar 

  31. 31.

    Eggleton, P. Separation of cells using free flow electrophoresis. in: Cell Separation. A Practical Approach. (Fisher, D., Francis, G.E. and Rickwood, D., Ed.) Oxford University Press, Oxford, New York, Tokyo 1998, 213–252.

    Google Scholar 

  32. 32.

    Hansen, E. Preparative free flow electrophoresis of lymphoid cells: A review. in: Cell Electrophoresis (Schütt, W., Klinkmann, H., Ed.) Walter de Gruyter, Berlin (NY) 1985, 287–304.

    Google Scholar 

  33. 33.

    Kuhn, R., Wagner, H., Mosher, R.A. and Thormann, W. Experimental and theoretical investigation of the stability of stepwise pH gradients in continuous flow electrophoresis. Electrophoresis 8 (1987) 503–508.

    CAS  Article  Google Scholar 

  34. 34.

    Wallach, D.F.H. and Lin, P.S. Plasma membrane fractionation. Biochim. Biophys. Acta 300 (1973) 211–254.

    PubMed  CAS  Google Scholar 

  35. 35.

    Morré, D.J., Morré, D.M. and van Alstine, J.M. Separation of endosomes by aqueous two-phase partition and free-flow electrophoresis. J. Chromatogr. B 711 (1998) 203–215.

    Google Scholar 

  36. 36.

    Toner, M. and Irimia, D. Blood-on-a-chip. Annu. Rev. Biomed. Eng. 7 (2005) 77–103.

    PubMed  CAS  Article  Google Scholar 

  37. 37.

    Barshtein, G., Tamir, I. and Yedgar, S. Red blood cell rouleaux formation in dextran solution: dependence on polymer conformation. Eur. Biophys. J. 27 (1998) 177–181.

    PubMed  CAS  Article  Google Scholar 

  38. 38.

    Bäumler, H., Donath, E., Krabi, A., Knippel, W., Budde, A. and Kiesewetter, H. Electrophoresis of human red blood cells and platelets. Evidence for depletion of dextran. Biorheology 33 (1996) 333–351.

    PubMed  Article  Google Scholar 

  39. 39.

    Gardner, B. The effect of dextrans on zeta potential. Proc. Soc. Exp. Biol. Med. 131 (1969) 1115–1118.

    PubMed  CAS  Google Scholar 

  40. 40.

    Ichiki, T., Ujiie, T., Shinbashi, S., Okuda, T. and Horiike, Y. Immunoelectrophoresis of red blood cells performed on microcapillary chips. Electrophoresis 23 (2002) 2029–2034.

    PubMed  CAS  Article  Google Scholar 

  41. 41.

    Lu, W.H., Deng, W.H., Liu, S.T., Chen, T.B. and Ra, P.F. Capillary electrophoresis of erythrocytes. Anal. Biochem. 314 (2003) 194–198.

    PubMed  CAS  Article  Google Scholar 

  42. 42.

    Omasu, F., Nakano, Y. and Ichik, T. Measurement of the electrophoretic mobility of sheep erythrocytes using microcapillary chips. Electrophoresis 26 (2005) 1163–1167.

    PubMed  CAS  Article  Google Scholar 

  43. 43.

    Walter, H. and Widen, K.E. Differential electrophoretic behavior in aqueous polymer solutions of red blood cells from Alzheimer patients and from normal individuals. Biochim. Biophys. Acta 1234 (1995) 184–190.

    PubMed  Article  Google Scholar 

  44. 44.

    Bäumler, H., Neu, B., Donath, E. and Kiesewetter, H. Basic phenomena of red blood cell rouleaux formation. Biorheology 36 (1999) 439–442.

    PubMed  Google Scholar 

  45. 45.

    Schüt, W., Thomaneck, U., Knippel, E., Rychly, J. and Klinkmann, H. Suitability of automated single cell electrophoresis (ASCE) for biomedical and clinical applications: General remarks. in: Cell Electrophoresis (Schütt, W., Klinkmann, H., Ed.) Walter de Gruyter, Berlin (NY) 1985, 313–332.

    Google Scholar 

  46. 46.

    Seaman, G.V.F. Electrokinetic behavior of red cells. in: The Red Blood Cells vol. 2. (Mac, D., Surgenor, N., Ed.), Academic Press, New York 1975, 1–135.

    Google Scholar 

  47. 47.

    Slivinsky, G.G., Hymer, W.C., Bauer, J. and Morrison, D.R. Cellular electrophoretic mobility data: A first approach to a database. Electrophoresis 18 (1997) 1109–1119.

    PubMed  CAS  Article  Google Scholar 

  48. 48.

    Josefowicz, J.Y. Electrophoretic light scattering and its application to the study of cells. in: Methods of Cell Separation, vol. 2. (Catsimpoolas, N., Ed.) Plenum Press NY 1979, 67–91.

    Google Scholar 

  49. 49.

    Walter, H. Cell partitioning in two-polymer aqueous phase systems. TIBS (1978) 97-100.

  50. 50.

    Hannig, K., Kowalski, M., Klock, G., Zimmermann, U. and Mang, V. Free-flow electrophoresis under microgravity: evidence for enhanced resolution of cell separation. Electrophoresis 11 (1990) 600–604.

    PubMed  CAS  Article  Google Scholar 

  51. 51.

    Todd, P. Microgravity cell electrophoresis experiments on the space shuttle: a 1984 overview. in: Cell Electrophoresis. (Schütt, W., Klinkmann H., Ed.), Walter de Gruyter, Berlin (NY) 1985, 3–19.

    Google Scholar 

  52. 52.

    Patel, D., Ford, T.C. and Rickwood, D. Fractionation of cells by sedimentation methods. in: Cell Separation. A Practical Approach. (Fisher, D., Francis, G.E. and Rickwood, D., Ed.) Oxford University Press, Oxford, New York, Tokyo 1998, 43–89.

    Google Scholar 

  53. 53.

    Malström, P., Nelson, K., Jönsson, A., Sjögren, H.O., Walter, H. and Albertsson, P.A. Separation of rat leukocytes by countercurrent distribution in aqueous two-phase systems. Cell Immunol. 37 (1978) 409–421.

    Article  Google Scholar 

  54. 54.

    Arnold, K., Herrmann, A., Pratsch, L. and Gawrisch, K. The dielectric properties of aqueous solutions of poly(ethylene glycol) and their influence on membrane structure. Biochim. Biophys. Acta 815 (1985) 515–518.

    PubMed  CAS  Article  Google Scholar 

  55. 55.

    Hansen, P.L., Cohen, J.A., Podgornik, R. and Parsegian, V.A. Osmotic properties of poly(ethylene glycols): quantitative features of brush and bulk scaling lows. Biophys. J. 84 (2003) 350–355.

    PubMed  CAS  Article  Google Scholar 

  56. 56.

    Sabolovic, D., Sestier, C., Perrotin, P., Guillet, R., Tefi, M. and Boynard, M. Covalent binding of polyethylene glycol to the surface of red blood cells as detected and followed up by cell electrophoresis and rheological methods. Electrophoresis 21 (2000) 301–306.

    PubMed  CAS  Article  Google Scholar 

  57. 57.

    Higuchi, A., Yamamiya, S., Yoon, B.O., Sakurai, M. and Hara, M. Peripheral blood cell separation through surface modified polyurethane membranes. J. Biomed. Materials Res. A 68A (2004) 34–42.

    CAS  Article  Google Scholar 

  58. 58.

    Di Basio, A. and Cametti, C. Effect of the shape of human erythrocytes on the evaluation of the passive electrical properties of the cell membrane. Bioelectrochemistry 65 (2005) 163–169.

    Article  CAS  Google Scholar 

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Correspondence to Włodzimierz Korohoda.

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Wilk, A., Rośkowicz, K. & Korohoda, W. A new method for the preperative and analytical electrophoresis of cells. Cell Mol Biol Lett 11, 579–593 (2006).

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Key words

  • Cell electrophoresis
  • Cell separation
  • Cell surface
  • Red blood cells