Integration of conventional cell viability assays for reliable and reproducible read-outs: experimental evidence

Abstract

ObjectiveShort-term viability assays of cultured cells in 96-well plates are routinely used to determine the cytotoxicity or safety of drugs. These are often based on the formation of chromogen, generated selectively in viable cells. The innate problems of such short-term cell viability assays include (i) effect of drugs is determined by cell density (ii) some drugs have slow/gradual effect and hence may escape such assays, (iii) cell morphology that reveal significant hints to molecular signaling underlining the effect of drugs cannot be effectively captured, (iv) long-term effect on viability and clonogenic potential of cells cannot be determined and (v) herbal extracts often possess intrinsic color that interferes with spectrophotometer estimation. In light of the ease and importance of cell culture-based assessment of drug safety and cytotoxicity, we attempted to combine the conventional cell-based assays in a way that allows multiple readouts (quantitative and qualitative) from a single experiment, and avoids the drawbacks of color interference.ResultsWe have established and validated (using 16 types of cultured mammalian cells) a Quantitative and Qualitative Cell Viability assay in 12-well cell culture plates. It overcomes several shortcomings as discussed above and allows long-term observations on cell morphology and clonogenicity.Electronic supplementary materialThe online version of this article (10.1186/s13104-018-3512-5) contains supplementary material, which is available to authorized users.

Identification and Functional Characterization of Anti-metastasis and Anti-angiogenic Activities of Triethylene Glycol Derivatives

“…The control or drug treated cells were incubated at 37°C and 5% CO 2 . After 48 h, cells were fixed, stained and de-stained into the solution, which was quantified by the help of a photospectrometer as described previously ( 27 ).…”

Rat Glioma Cell-Based Functional Characterization of Anti-Stress and Protein Deaggregation Activities in the Marine Carotenoids, Astaxanthin and Fucoxanthin

“…Based on these data, 2.5 µM Asta (A1) and 0.25 µM Fuco (F1) were chosen as the nontoxic doses. Both A1 and F1 treated cells showed no significant difference, with respect to control cells, both on short- and long-term viability, clonogenicity as well as cell morphology as observed by a quantitative cell viability assay [2] and cell proliferation assay, as shown in Figure 1C,D. Therefore, these nontoxic doses were used for further experiments.…”

“…The drug-supplemented medium was replaced every alternate day. After 8 days, cells were fixed, stained, and de-stained into the solution, which was quantified by the help of a spectrophotometer as described previously [2]. Statistical significance was calculated by an unpaired t -test of GraphPad ® software (2018) using mean, SD, and N from three independent experiments, and shown as * p < 0.05, ** p < 0.01, *** p < 0.001, ns = not significant.…”

“…On these premises, there is a continuous search and enrollment of natural compounds with anti-stress potential for extending quality of life in normal and disease scenarios. Cell-based functional assays provide a useful model system to identify such physiologically relevant compounds [2]. Oxidative and UV-induced stresses are some of the most common stresses that cause molecular damage both at the DNA and protein level [3], and are often linked to the lethal pathologies including cancer and brain dysfunctions [1,4].…”

Optimization of Culture Media for Halocnemum strobilaceum Associated Endophytes Bioactivity

“…Screening in-vitro cytotoxicity and viability on cellular parameters using MTT (3-[4,5dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay as a fast, cheap, and highly sensitive method [6]. Breast cancer cell line (MCF- 7) retained mammary epithelium characteristics useful for experimental therapeutics to determine the cytotoxicity or safety of drugs [7,8].…”