Abstract
Apparatus and methods were developed to enable mechanically facilitated cell-cell electrofusion to be performed. The apparatus and methods mechanically place cells in contact before fusion. The key component of this fusion system was a newly developed fusion chamber. The chamber was composed of two functionally identical electrodes that were housed in a multi-layer structure. The layers functioned as support for the electrodes. They also allowed adjustment of the distance between opposing electrode faces. The electrodes were constructed in a manner that allowed cells to be deposited, by vacuum, onto each face. Electrode faces were positioned at a predetermined distance from each other to mechanically force cell-cell contact between the deposited cells. Fusion was induced by delivering direct current pulses to the juxtaposed cells. Fusion products were detected and quantitated by flow cytometry. Details of the chamber design and a protocol for using the fusion chamber are given. Mechanically facilitated cell-cell electrofusion was demonstrated by using the chamber to produce fusion products from like fusion partners. The practical applicability of the chamber was demonstrated by fusing unlike cell types. Mechanically facilitated cell-cell electrofusion is not specific to the cells used in this study; the chamber can be adapted for use with other cell types.
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- Abidor I. G., Sowers A. E. Kinetics and mechanism of cell membrane electrofusion. Biophys J. 1992 Jun;61(6):1557–1569. doi: 10.1016/S0006-3495(92)81960-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bakker Schut T. C., Kraan Y. M., Barlag W., de Leij L., de Grooth B. G., Greve J. Selective electrofusion of conjugated cells in flow. Biophys J. 1993 Aug;65(2):568–572. doi: 10.1016/S0006-3495(93)81128-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Dimitrov D. S., Apostolova M. A., Sowers A. E. Attraction, deformation and contact of membranes induced by low frequency electric fields. Biochim Biophys Acta. 1990 Apr 30;1023(3):389–397. doi: 10.1016/0005-2736(90)90131-7. [DOI] [PubMed] [Google Scholar]
- Finaz C., Lefevre A., Teissié J. Electrofusion. A new, highly efficient technique for generating somatic cell hybrids. Exp Cell Res. 1984 Feb;150(2):477–482. doi: 10.1016/0014-4827(84)90592-5. [DOI] [PubMed] [Google Scholar]
- Foung S. K., Perkins S. Electric field-induced cell fusion and human monoclonal antibodies. J Immunol Methods. 1989 Jan 6;116(1):117–122. doi: 10.1016/0022-1759(89)90319-0. [DOI] [PubMed] [Google Scholar]
- Glassy M. Creating hybridomas by electrofusion. Nature. 1988 Jun 9;333(6173):579–580. doi: 10.1038/333579a0. [DOI] [PubMed] [Google Scholar]
- Grasso R. J., Heller R., Cooley J. C., Haller E. M. Electrofusion of individual animal cells directly to intact corneal epithelial tissue. Biochim Biophys Acta. 1989 Mar 27;980(1):9–14. doi: 10.1016/0005-2736(89)90193-4. [DOI] [PubMed] [Google Scholar]
- Heller R., Grasso R. J. Transfer of human membrane surface components by incorporating human cells into intact animal tissue by cell-tissue electrofusion in vivo. Biochim Biophys Acta. 1990 May 9;1024(1):185–188. doi: 10.1016/0005-2736(90)90223-b. [DOI] [PubMed] [Google Scholar]
- Hewish D. R., Werkmeister J. A. The use of an electroporation apparatus for the production of murine hybridomas. J Immunol Methods. 1989 Jun 21;120(2):285–289. doi: 10.1016/0022-1759(89)90254-8. [DOI] [PubMed] [Google Scholar]
- Jaroszeski M. J., Gilbert R., Heller R. Detection and quantitation of cell-cell electrofusion products by flow cytometry. Anal Biochem. 1994 Feb 1;216(2):271–275. doi: 10.1006/abio.1994.1041. [DOI] [PubMed] [Google Scholar]
- Lo M. M., Tsong T. Y., Conrad M. K., Strittmatter S. M., Hester L. D., Snyder S. H. Monoclonal antibody production by receptor-mediated electrically induced cell fusion. 1984 Aug 30-Sep 5Nature. 310(5980):792–794. doi: 10.1038/310792a0. [DOI] [PubMed] [Google Scholar]
- Lojewska Z., Farkas D. L., Ehrenberg B., Loew L. M. Analysis of the effect of medium and membrane conductance on the amplitude and kinetics of membrane potentials induced by externally applied electric fields. Biophys J. 1989 Jul;56(1):121–128. doi: 10.1016/S0006-3495(89)82657-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Marszalek P., Liu D. S., Tsong T. Y. Schwan equation and transmembrane potential induced by alternating electric field. Biophys J. 1990 Oct;58(4):1053–1058. doi: 10.1016/S0006-3495(90)82447-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Montané M. H., Dupille E., Alibert G., Teissié J. Induction of a long-lived fusogenic state in viable plant protoplasts permeabilized by electric fields. Biochim Biophys Acta. 1990 May 9;1024(1):203–207. doi: 10.1016/0005-2736(90)90227-f. [DOI] [PubMed] [Google Scholar]
- Pohl H. A., Crane J. S. Dielectrophoresis of cells. Biophys J. 1971 Sep;11(9):711–727. doi: 10.1016/S0006-3495(71)86249-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sowers A. E. A long-lived fusogenic state is induced in erythrocyte ghosts by electric pulses. J Cell Biol. 1986 Apr;102(4):1358–1362. doi: 10.1083/jcb.102.4.1358. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sowers A. E. Fusion events and nonfusion contents mixing events induced in erythrocyte ghosts by an electric pulse. Biophys J. 1988 Oct;54(4):619–626. doi: 10.1016/S0006-3495(88)82997-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sukharev S. I., Bandrina I. N., Barbul A. I., Fedorova L. I., Abidor I. G., Zelenin A. V. Electrofusion of fibroblasts on the porous membrane. Biochim Biophys Acta. 1990 May 16;1034(2):125–131. doi: 10.1016/0304-4165(90)90065-5. [DOI] [PubMed] [Google Scholar]
- Teissie J., Blangero C. Direct experimental evidence of the vectorial character of the interaction between electric pulses and cells in cell electrofusion. Biochim Biophys Acta. 1984 Sep 5;775(3):446–448. doi: 10.1016/0005-2736(84)90203-7. [DOI] [PubMed] [Google Scholar]
- Teissie J., Knutson V. P., Tsong T. Y., Lane M. D. Electric pulse-induced fusion of 3T3 cells in monolayer culture. Science. 1982 Apr 30;216(4545):537–538. doi: 10.1126/science.7071601. [DOI] [PubMed] [Google Scholar]
- Teissie J., Rols M. P. Fusion of mammalian cells in culture is obtained by creating the contact between cells after their electropermeabilization. Biochem Biophys Res Commun. 1986 Oct 15;140(1):258–266. doi: 10.1016/0006-291x(86)91084-3. [DOI] [PubMed] [Google Scholar]
- Wojchowski D. M., Sytkowski A. J. Hybridoma production by simplified avidin-mediated electrofusion. J Immunol Methods. 1986 Jun 24;90(2):173–177. doi: 10.1016/0022-1759(86)90073-6. [DOI] [PubMed] [Google Scholar]
- Zimmermann U. Electric field-mediated fusion and related electrical phenomena. Biochim Biophys Acta. 1982 Nov 30;694(3):227–277. doi: 10.1016/0304-4157(82)90007-7. [DOI] [PubMed] [Google Scholar]



