Study On The Electrochemical Method For Detection Of Cell Activity

Objective In this study,Balb/c mouse embryonic fibroblasts?Balb/c 3T3?,Chinese hamster ovary?CHO?,rat adrenal pheochromoma?PC-12?and Chinese hamster lung fibroblasts?V79?were used as cell models to study the electrochemical behavior of four kinds of cells,and the characteristic of cell activity and cytotoxicity in electrochemical detection process by using purines as biomarkers.The theoretical and experimental basis was provided to establish a simple,rapid,sensitive,low-cost,lable-free electrochemical assay for cell activity.Methods The electrochemical behaviors of Balb/c 3T3,CHO,PC-12 and V79 cells were studied by cyclic voltammetry.Three electrode systems with multi-walled carbon nanotubes modified glassy carbon electrode?C-MWCNTs/GCE?as working electrode,Ag/AgCl as reference electrode and Pt as counter electrode were used for electrochemical detection.The oxidation peaks of four cell suspensions were assigned by enzymatic catalysis method and standard comparison method.The four different cell growth curves,the effect of serum concentration on cell activity,the influences of drug-treated time and drug concentration on cytotoxicity were studied by electrochemistry,cell counting,3-?4,5-dimethylthiazole-2?-2,5-diphenyltetrazolium bromide?MTT?and trypan blue staining methods.The results revealed the characteristics using purine as the bio-markers in reflection of cell activity and toxicity.Results Three oxidation peaks assigned as uric acid?0.31 V?,xanthine/guanine?0.69V?and hypoxanthine?1.01 V?in Balb/c 3T3,PC-12 and V79 cell suspensions,and assigned as xanthine/guanine?0.69 V?,adenine?0.95 V?and hypoxanthine?1.01 V?in CHO cell suspensions were detected,respectively.The good linear relationships between mixed oxidation peak current values of xanthine/guanine and cell concentrations in the range from 1.25×10⁵ to 80×10⁵ Cells/mL were detected,and the linear relations were y=0.165x+0.118?R²=0.994?,y=0.124x+0.179?R²=0.981?,y=0.130x+0.442?R²=0.985?and y=0.118x+0.200?R²=0.990?,respectively.The general trends of four different cells growth curves plotted by electrochemical and cell counting method were consistent,which showed change characteristics in lag,logarithmic,plateau and decay periods.The effects of serum concentration on cell viability detected by electrochemical method were consistent with that detected by MTT and cell counting.The relationships between cytotoxicity of N-methyl-N'-nitro-N-nitrosoguanidine?MNNG?and exposure time,and concentration were detected by electrochemical method.The general trend detected by electrochemical method was consistent with MTT,cell counting and trypan blue staining.Compared with MTT method,the cytotoxic platform of drugs could be detected 12 hours in advance by electrochemical method,and the IC₅₀ values calculated by electrochemical method were less than MTT,cell counting and trypan blue exclusion method,which reflected its sensitivity in detection of cytotoxicity.In addition,the inhibitory effect of drug concentration on the growth of cells in a dose-dependent manner could be reflected by the electrochemical method.Conclusions With purines as biomarkers,the same trends of different cell lines were detected by electrochemical method in cell viability and cytotoxicity.Electrochemistry exhibited more sensitive characteristics than MTT,cell counting and trypan blue exclusion method.With purines contents as indicators,the changes of cell activity and cytotoxicity can be quickly,sensitively and accurately detected by the electrochemical detection of cell activity.