Measurement of cytotoxicity by ATP-based luminescence assay in primary cell cultures and cell lines.
Drug discovery and toxicological safety testing share a need for dependable in vitro cellular toxicity tests. Ideally such tests should be objective, quantitative, reproducible and able to lend themselves to automation. A number of assays fulfil these criteria well, but recently it has become clear that the molecular phenotype of the cell tested and the complex interplay between different cell types can radically alter the response to individual agents. The differences observed between primary cell cultures and cell lines make it preferable to use primary cultures for assessment of toxicity, yet the problems of using primary cell cultures are considerable as the number of cells available for testing is often small. Recently, we have developed a short-term cell culture assay based on the detection of ATP by the luciferin-luciferase reaction. Four drugs/agents can be tested in triplicate at seven dilutions in one 96-well microplate with 1000 cells/well in the case of cell lines, or 10,000 cells/well for primary tumour tissue. The small number of cells required is a major advantage of this method. Initially developed as a tumour chemosensitivity assay, the assay has shown considerable promise as a general in vitro toxicity assay allowing both cell lines and primary tissue cultures to be tested. Heterogeneity of sensitivity is present in benign tissue biopsies as well as tumours. Molecular alterations within the cell and the interplay of different cell types have been addressed in a number of different model systems using the assay, suggesting that this technology may have more general application.
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Department of Pathology, Institute of Ophthalmology, University College London, Bath Street, London EC1V 9EL, UK.
Andreotti
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