Skip to main content
Log in

Comparison of 5 microplate colorimetric assays forin vitro cytotoxicity testing and cell proliferation assays

  • Published:
Cytotechnology Aims and scope Submit manuscript

Abstract

This paper describes a critical comparative evaluation of 5 miniaturised colorimetric assays applicable to cytotoxicity testing of anti-tumour drugs (and other toxins)in vitro. Each assay shows a different linear range for optical density versus cell number, a different sensitivity to change in cell number and a different minimum detectable cell number; the values of these parameters vary with experimental conditions and with cell line used. All the methods gave good correlation with viable cell number (determined by colony forming efficiency) in toxicity assays after 3 or 4 days of treatment, but they underestimated cell death after 2 days. Toxicity levels for individual chemicals (in a standard 6-day assay) are similar for the different assays, irrespective of the mechanism of action of the chemical being tested. Two of the more recently developed assays (APNaOH and SRB) were found to be very sensitive under the conditions examined.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Similar content being viewed by others

References

  • Alley MC, Scudiero DA, Monks A, Hursey ML, Czerwinsk MJ, Fine DL, Abbott BJ, Mayo JG, Shoemaker RH and Boyd MR (1988) Feasibility of drug screening with panels of human tumor cell lines using a microculture tetrazolium assay. Cancer Res. 48: 589–601.

    Google Scholar 

  • Borenfreund E and Puemer JA (1983) A simple quantitative procedure using monolayer cultures for cytotoxicity assays. J. Tissue Culture Meth. 9: 7–9.

    Google Scholar 

  • Connolly DT, Knight MB, Harakas NK, Wittwer AJ and Feder J (1986) Determination of the number of endothelial cells in culture versus an acid phosphatase assay. Anal. Biochem. 152: 136–140.

    Google Scholar 

  • Cook JA and Mitchell JB (1989) Viability measurements in mammalian cell systems. Anal. Biochem. 179: 1–7.

    Google Scholar 

  • Fiennes A, Walton J, Winterboume D, McGlashan D and Hermon-Taylor J (1987) Quantitative correlation of natural red dye uptake with cell numbers in human cancer cell cultures. Cell Biol. Int. Repts. 11: 373–378.

    Google Scholar 

  • Gilvarry U, Farrell D, Lynch V, Moriarty M, Dooley M, Law E and Clynes M (1990) Cytological differences between normal and malignant cell populations in culture. Cancer Res. 50, 3390–3393.

    Google Scholar 

  • Givens K, Kirada S, Chen A, Rothschiller J and Lee D (1990) Purification of human ocular fibroblasts. An assessment ofin vitro colorimetric assays. Invest. Ophthalmol. Vis. Sci. 31: 1856–1862.

    Google Scholar 

  • Huschtscha LI, Lucibello FC and Bodmer WF (1989) A rapid method for counting cells ‘in situ’ using a fluorogenic alkaline phosphatase enzyme assay. In Vitro Cell Dev. Biol. 25: 105–108.

    Google Scholar 

  • Keepers YP, Pizao FE, Peters GJ, Vanarkotte J, Winograd B and Pinedo HM (1991) Comparison of the sulforhodamine-B protein and tetrazolium (MTT) assays forin vitro chemosensitivity testing. Eur. J. Cancer 27: 897–900.

    Google Scholar 

  • Kueng W, Silber E and Eppenberger U (1989) Quantification of cells cultured on 96-well plates. Anal. Biochem. 182: 16–19.

    Google Scholar 

  • Martin A and Clynes M (1991) Acid phosphatase: endpoint forin vitro toxicity tests. In Vitro Cell Dev. Biol. 27A: 183–184.

    Google Scholar 

  • Miura M, Kunitomo K and Komi N (1988)In vitro chemosensitivity testing based on intracellular ATF levels (ATP assay). Tokushima. J. Exp. Med 35: 49–55.

    Google Scholar 

  • Mossman T (1983) Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J. Immunol. Meth. 65: 55–63.

    Google Scholar 

  • Neville ME (1987)51Cr uptake assay: a sensitive and reliable method to quantitate cell viability and cell death. J. Immunol. Meth. 99: 77–82.

    Google Scholar 

  • Scragg M and Ferreira L (1991) Evaluation of differential staining procedures for quantification of fibroblasts cultured in 96-well plates. Anal. Biochem. 198: 80–85.

    Google Scholar 

  • Skehan F, Storeng R, Scudiero D, Monks A, McMahon J, Vistico D, Warren J, Bokesch H, Kenney S and Boyd MR (1990) New colorimetric assay of anticancer drug screening. J. Natl. Canc. Inst. 82: 1107–1113.

    Google Scholar 

  • Triglia D, Braa S, Yonan C and Naughton G (1991)In vitro toxicity of various classes of test agents using the neutral red assay on a human three-dimensional physiologic skin model. In Vitro Cell Dev. Biol. 27A: 239–244.

    Google Scholar 

  • Vistica DT, Skehan P, Scudiero D, Monks A, Pittman A and Boyd MR (1991) Tetrazolium-based assays for cellular viability: a critical examination of selected parameters affecting formazan production. Cancer Res. 51: 2515–2520.

    Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Rights and permissions

Reprints and permissions

About this article

Cite this article

Martin, A., Clynes, M. Comparison of 5 microplate colorimetric assays forin vitro cytotoxicity testing and cell proliferation assays. Cytotechnology 11, 49–58 (1993). https://doi.org/10.1007/BF00749057

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00749057

Key words

Navigation