Cell-cell electrofusion: optimization of electric field amplitude and hypotonic treatment for mouse melanoma (B16-F1) and Chinese Hamster ovary (CHO) cells.
Efficient electroporation of cells in physical contact induces cell fusion, and this process is known as electrofusion. It has been shown that appropriate hypotonic treatment of cells before the application of electric pulses can cause a significant increase in electrofusion efficiency. First, the amplitudes of the electric field were determined spectrofluorometrically, where sufficient permeabilization in hypotonic buffer occurred for B16-F1 and CHO cells. In further electrofusion experiments 14 +/- 4% of fused cells for B16-F1 and 6 +/- 1% for CHO was achieved. These electrofusion efficiencies, determined by double staining and fluorescence microcopy, are comparable to those of other published studies. It was also confirmed that successful electroporation does not necessarily guarantee high electrofusion efficiency due to biological factors involved in the electrofusion process. Furthermore, not only the extension of electrofusion but also cell survival depends on the cell line used. Further studies are needed to improve overall cell survival after electroporation in hypotonic buffer, which was significantly reduced, especially for B16-F1 cells. Another contribution of this report is the description of a simple modification of the adherence method for formation of spontaneous cell contact, while cells preserve their spherical shape.
Usaj
Marko
M
Faculty of Electrical Engineering, University of Ljubljana, Trzaska 25, 1000, Ljubljana, Slovenia.
Trontelj
Katja
K
Miklavcic
Damijan
D
Kanduser
Masa
M
D000818:Animals; D016466:CHO Cells; D002459:Cell Fusion / Q000379:methods*; D045744:Cell Line, Tumor; D002470:Cell Survival; D006224:Cricetinae; D003412:Cricetulus; D055664:Electrochemical Techniques; D051379:Mice; D008856:Microscopy, Fluorescence / Q000379:methods