Cytotoxicity assessment based on the AUC50 using multi-concentration time-dependent cellular response curves

• Published in: Analytica chimica acta 2013-02, Vol.764, p.44-52
• Main Authors: Pan, Tianhong, Huang, Biao, Zhang, Weiping, Gabos, Stephan, Huang, Dorothy Yu, Devendran, Vignesh
• Format: Article
• Language: English
• Subjects: Area Under Curve; Area under the control line; Cell Survival - drug effects; Colchicine - toxicity; Hep G2 Cells; High-throughput screening; Humans; In vitro cytotoxicity assay; Models, Theoretical; Repeatability; Time Factors; Time-dependent cellular response curve; Toxicity Tests
• ISSN: 0003-2670; 1873-4324
• DOI: 10.1016/j.aca.2012.12.047

[Display omitted] ► Dose- and time-dependent cellular responses are used to evaluate the cytotoxicity. ► The CI can reflect the cell number, cell viability and morphological change, etc. ► AUC is more relevant to the intensity of the cell treatment. ► AUC50 can be used for cytotoxicity assessment. ► AUC50 combined with RTCA HT assay can achieve a high-throughput screening. Although many indices have been developed to quantify chemical toxicity, substantial shortcoming is inherent in most of them, such as observation time dependence, insufficient robustness, and no comparison with the negative control. To assess the extent of exposure of the tested substance, a cytotoxicity assay named AUC50 was developed to describe the time and concentration-dependent cellular responses. By monitoring the dynamic cytotoxicity response profile of living cells via the xCELLigence real-time cell analysis high-throughput (RTCA HT) system, changes in cell number (named cell index, CI) were recorded and analyzed subsequently. A normalized cell index (NCI) is introduced to reduce the influence of inter-experimental variations. The log-phase of cellular growth is considered, which alleviates the cell's spontaneous effect. The area between the control line and the assessed time-dependent cellular response curve (TCRC) of the tested substance was calculated, and the corresponding exponential kill model (concentration–response curve) was developed along with exploiting the concept of AUC50. The validation of the proposed method is demonstrated by exposing HepG2 cell line to seven chemical compounds. Our findings suggested that the proposed AUC-based toxicity assay could be an alternative to the traditional single time-point assay, and it has potential to become routine settings for evaluating the cell-based in vitro assay. Furthermore, the AUC50 combined with RTCA HT assay can be used to achieve a high-throughput screening that conventional cellular assay cannot achieve.