Presentation on theme: "SEMINAR-III 28-09-09 SPEAKER KALPANA. Impedimetric monitoring of cell attachment on interdigitated microelectrodes Bin-Wha Chang a,b, Che-Hsiung Chena,"— Presentation transcript:
SEMINAR-III SPEAKER KALPANA
Impedimetric monitoring of cell attachment on interdigitated microelectrodes Bin-Wha Chang a,b, Che-Hsiung Chena, Shin-Jyh Ding c, David Chan-Hen Chend, Hsien-Chang Changa, ∗ a Institute of Biomedical Engineering, National Cheng-Kung University, Tainan 701, Taiwan, ROC b Department of Healthcare Administration, Hung-Kuang University, Taichung 433, Taiwan, ROC c Institute of Dental Materials, Chung-Shan Medical University, Taichung 402, Taiwan, ROC d Institute of Veterinary Microbiology, National Chung-Hsing University, Taichung 402, Taiwan, ROC
1.INTRODUCTION Cell attachment is the initial step in tissue or tumor formation process including cell growth, migration, junction formation, metabolism, division, differentiation, metastasis and apoptosis. To assay cell attachment, direct cell counting, time-lapse cinematograph and colorimetric methods have conventionally been used. –laborious, indirect and not time-resolved. Ehret et al. designed an interdigitated microelectrode to obtain the impedance data attributable to cell physiological status, the interdigitated electrode design has merits of high cell constant and uniform sensing interval. –very reliable and informative.
2.EXPERIMENTAL Schematic diagram of the impedimetric system:
2.1. Impedimetric measuring chamber with microelectrodes Interdigitated microelectrode plate (IMP, Fig. 1c) was patterned on a glass slide (26mm × 76mm; Fig. 1b) by the wet-etching process. Cell chamber for impedimetric measurement was constructed by mounting and fixing a section of glass tube (12mm i.d. × 6 cm height) on the IMP plate using epoxy resin. (b) schematic diagram of the interdigitated microelectrode plate; (c) a photograph of MDCK cells grown on the microelectrode plate.
RGD-C peptide was adsorbed on the microelectrodes 1 mg/cm2 by dipped-coating procedure Culturing cells in the measuring chamber MDCK cells (ATCC CCL-34, passages 50–90) were cultured in Dulbeco’s modified Eagle’s medium (DMEM) supplemented with 10% fetalbovine serum (GIBCO) and 5% gentamycin at 37 ◦C in a humidified atmosphere of 5% CO2 and 95% air Impedance analyzing system Time-resolved admittance spectra were obtained with a commercialized impedance analyzer (hp4194A). The operating frequency was scanned from 1 kHz to 1MHz by 10 kHz increment, and the sampling interval was 1 min.
3. Results and discussion Monitoring of admittance during cell attachment:
`Determination of cell density using low-frequency conductance signal:
The following correlation equation was established from 30 experiments (Fig. 4) with different cell densities using chambers with bare microelectrodes: The following equation was established using chambers with electrodes coated with RGD-C peptide (n = 20; Fig. 4):
4. Conclusions Data obtained with the impedimetric system were informative for cell physiology and biomaterial studies. Cell attaching/detaching process, formation of confluent layer, apoptosis and even migration phenomena might be resolved and quantified by the proposed system. The study provided also a reliable protocol to determine the cell density attached onto the surface interested and also the timing of the attach process.