Cell reactions to dielectrophoretic manipulation.
The phenomenon of dielectrophoretic particle manipulation holds promise for many biotechnology applications, including cell sorting. In our system cell manipulation normally involves transient exposure (15 minutes) to radio-frequency AC electric fields generated using planar microelectrodes. The present study was designed to investigate the range of acute effects of dielectrophoretic manipulation on the normal physiology of isolated cells. Cells were suspended in isoosmotic Mannitol and exposed to a 5 MHz, 21 V (peak to peak) electric field with 100 micrometer gap electrodes. Cells were assigned to three experimental groups; non-exposed controls, exposed cells processed immediately after cessation of the field, and exposed cells processed after a time delay. SEM observations of spread cells cultured on the devices showed no apparent acute effects of field exposure on cell morphology. Cell-doubling rates in exposed cells subsequent to field-exposure or transient incubation in mannitol were no different from control cells. An MTT 'mitochondrial stress' assay indicated no alteration in the rate of oxidative respiration in exposed cells 0.5 hour after exposure to the field. Western blot analysis indicated upregulation of fos protein in cells 0.5 hour after field-exposure, which was confirmed using densitometry. Reverse transcription of cellular mRNA followed by PCR amplification, polyacrylamide gel electrophoresis and autoradiography of cDNA banding revealed differential gene expression between controls and exposed cells processed immediately after cessation of the field. Differential gene expression persisted in exposed cells at least 0.5 hours after removal from the field. Observations indicated that temperature fluctuation in the mannitol solution was minimal, suggesting that upregulated mRNA may not have been related to thermally-induced heat shock protein. The present study has indicated that exposure to AC fields during dielectrophoretic cell manipulation is associated with upregulation of the intermediate-early gene cfos and also transcription of other as yet unidentified genes. These transcriptional events were not manifest as gross changes in cell morphology or cell-cycle dynamics.
Archer
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Department of Electronics and Electrical Engineering, University of Glasgow, Glasgow, G12 8QQ, United Kingdom.
Li
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Evans
A T
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Britland
S T
ST
Morgan
H
H
D000818:Animals; D015153:Blotting, Western; D002455:Cell Division; D002460:Cell Line; D002468:Cell Physiological Phenomena; D019069:Cell Respiration; D048429:Cell Size; D004586:Electrophoresis / Q000295:instrumentation / Q000379:methods; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D015870:Gene Expression; D008353:Mannitol / Q000378:metabolism; D008564:Membrane Potentials; D008839:Microelectrodes; D008928:Mitochondria / Q000502:physiology; D009994:Osmolar Concentration; D011209:Power, Psychological; D016760:Proto-Oncogene Proteins c-fos / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D013696:Temperature; D013997:Time Factors; D015854:Up-Regulation
D016760:Proto-Oncogene Proteins c-fos; D012333:RNA, Messenger; D008353:Mannitol