title: A biophysical approach to the optimisation of dendritic-tumour cell electrofusion.
abstract: Electrofusion of tumour and dendritic cells (DCs) is a promising approach for production of DC-based anti-tumour vaccines. Although human DCs are well characterised immunologically, little is known about their biophysical properties, including dielectric and osmotic parameters, both of which are essential for the development of efficient electrofusion protocols. In the present study, human DCs from the peripheral blood along with a tumour cell line used as a model fusion partner were examined by means of time-resolved cell volumetry and electrorotation. Based on the biophysical cell data, the electrofusion protocol could be rapidly optimised with respect to the sugar composition of the fusion medium, duration of hypotonic treatment, frequency range for stable cell alignment, and field strengths of breakdown pulses triggering membrane fusion. The hypotonic electrofusion consistently gave a tumour-DC hybrid rate of up to 19%, as determined by counting dually labelled fluorescent hybrids in a microscope. This fusion rate is nearly twice as high as that usually reported in the literature for isotonic media. The experimental findings and biophysical approach presented here are generally useful for the development of efficient electrofusion protocols, especially for rare and valuable human cells.
authors.affiliation: Lehrstuhl für Biotechnologie, Biozentrum, Universität Würzburg, Am Hubland, D-97074 Würzburg, Germany.
authors.forename: Vladimir L
authors.initials: VL
authors.lastname: Sukhorukov
authors.forename: Randolph
authors.initials: R
authors.lastname: Reuss
authors.forename: Jörg M
authors.initials: JM
authors.lastname: Endter
authors.forename: Steffen
authors.initials: S
authors.lastname: Fehrmann
authors.forename: Alisa
authors.initials: A
authors.lastname: Katsen-Globa
authors.forename: Petra
authors.initials: P
authors.lastname: Gessner
authors.forename: Andrea
authors.initials: A
authors.lastname: Steinbach
authors.forename: Kilian J
authors.initials: KJ
authors.lastname: Müller
authors.forename: Abraham
authors.initials: A
authors.lastname: Karpas
authors.forename: Ulrich
authors.initials: U
authors.lastname: Zimmermann
authors.forename: Heiko
authors.initials: H
authors.lastname: Zimmermann
mesh_terms: D055592:Biophysical Phenomena; D001703:Biophysics; D002241:Carbohydrates; D002459:Cell Fusion; D048429:Cell Size; D002478:Cells, Cultured; D003713:Dendritic Cells / Q000166:cytology*; D004563:Electrochemistry; D006801:Humans; D007038:Hypotonic Solutions; D008855:Microscopy, Electron, Scanning; D009369:Neoplasms / Q000473:pathology*
chemical_list: D002241:Carbohydrates; D007038:Hypotonic Solutions