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Cell detachment using electric discharge plasmas Ingrid E. Kieft, Eva Stoffels, Veronique Caubet-Hilloutou, Jos L.V. Broers, Frans C.S. Ramaekers, Dick W. Slaaf Department of Biomedical Engineering, Eindhoven University of Technology P.O.Box 513, 5600 MB Eindhoven, The Netherlands E-mail: [email protected] Results and discussion Introduction • Study of interaction of small atmospheric discharge with living cells • Use of novel plasma source the “plasma needle” [1] • Effects studied on both CHO K1 and human epithelial NSCLC MR65 cells • Exposed cells detach instantaneously from the surface • Loss of cell-cell interaction • Detached cells remain alive and reattach within several hours to surface • Plasma treatment can be performed with high precision with spot-sizes smaller than 1 mm • Neighbouring cells are left unaffected • Treatment of cells with high dose can cause necrosis • If the right dose is applied, cells detach • A void can be created with dimensions sub-mm necrotic zone void where cells detached zone with rounded and living cells Treatment with high dose. Dead cells in red on top, then a void caused by detached cells, the green cells at the bottom are alive. Sharp boundaries! Material and methods Plasma generation •We ignite the RF plasma in helium. The frequency we use is about 13.2 MHz. •Plasma appears at the tip of a metal pin. It appears as a pink glow. On the mechanism •Helium: inert, has largest heat conductivity of noble gases •Plasma temperature: below 30oC •Plasma influence expected from radicals and ions •Cells shielded from UV light by a PBS solution •Generated electric field not affecting cells waveform generator RF amplifier Cell detachment in time. 1: control sample, 2:15 min. after treatment, 3: 1 hours after treatment, 4: 4 hours after treatment. power meter • • matching network • Cells start attaching one hour after treatment Same effect for fibroblasts and epithelial cells cell detachment is general effect Most likely cell adhesion molecules destroyed by plasma radicals [2] He bottle flow control unit Plasma discharge Scheme of the experimental setup Methods Plasma parameters •Plasma power is 50 to 100 mW •Voltage peak to peak 200 - 400 V •Helium flow 2 l/min •Total irradiation time of sample is 1 minute • individual cells are irradiated for 5 to 10 seconds Viability assays •PI and CTG are used to distinguish dead and living cells Proposed mechanism for detachment of cells. Conclusions • Cells detach after plasma treatment, and remain alive. They are capable of reattachment and cytokinesis. • The plasma treatment can be performed with sub-millimetre precision. Neighbouring cells are not affected. References: • The high precision plasma treatment can have applications in wound healing and cancer treatment. [1] Stoffels, E., Flikweert, A.J., Stoffels, W.W., Kroesen, G.M.W., 2002, Plasma Sources Sci. Technol. 11: 383-388 [2] Stoffels,E., Kieft, I.E., Sladek, R.E.J., Superficial treatment of mammalian cells using plasma needle, accepted for publication in J.Phys.D: Appl. phys.