Abstract
The artificially induced cell fusion is a useful experimental tool in biology, biotechnology and medicine. The electrofusion is a physical method for cell fusion that applies high-voltage electric pulses. The use of electric pulses causes cell membrane structural changes which bring the cell membrane in the so-called fusogenic state. When such fusogenic membranes are in close contact cell fusion takes place. Physical contact between fusion partners can be achieved by various methods and one of them is modified adherence method (MAM) described in detail here on B16-F1 cell line. The method is based on the fact that living cells form contacts in confluent culture. However, instead of using confluent cell culture, in modified adherence method cells are plated in suitable concentration and allowed to form contacts for only short predetermined period of time. During that time the cells are only slightly attached to the dish surface maintaining the spherical shape. Observed high fusion yields up to 50 % obtained by MAM in situ by dual-color fluorescence microscopy are among the highest in field of electrofusion. The method can be readily adapted to other anchorage-dependent cell lines.
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Acknowledgement
This work was supported by Infrastructure Program: Network of research infrastructure centers at University of Ljubljana (2009–2014) IP-0510 and Research program P2-0249, founded by Slovenian Research Agency (ARRS), Slovenia. Research was conducted in the scope of the LEA EBAM European Associated Laboratory (LEA). We would like to thank D. Miklavčič, the head of Laboratory of Biocybernetrics, for his general support and M. Simčič from Laboratory of Modeling, Simulation and Control, Faculty of Electrical Engineering, University of Ljubljana for the photo of the multiwell plate in the Fig. 1.
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Ušaj, M., Kandušer, M. (2015). Modified Adherence Method (MAM) for Electrofusion of Anchorage-Dependent Cells. In: Pfannkuche, K. (eds) Cell Fusion. Methods in Molecular Biology, vol 1313. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2703-6_15
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DOI: https://doi.org/10.1007/978-1-4939-2703-6_15
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