Miniaturization and application of the MTT assay to evaluate metabolic activity of protozoa in the presence of toxicants

This paper describes a critical evaluation of a miniaturised colorimetric a ssay, using MTT(3-[4,5-dimethyl-thiazol-2-yl] -2,5-diphenyl-tetrazolium bro mide) reduction, applied to protozoan viability testing. The toxic substanc es used were copper, zinc, Triton X-100 (a membrane surfactant) and cyclohe ximide (an inhibitor of the protein synthesis). The viability assay of the ciliate protozoan Tetrahymena pyriformis was optimised in terms of MTT conc entration and incubation time. Since protozoa are non adherent cells the MT T assay was modified in order to maintain the medium in the well. MTT prove d to be effective in the measurement of Tetrahymena pyriformis viability. F our hours of MTT incubation followed by 30 minutes of incubation with DMSO were found to be the best incubation times for optical density reading. Fur thermore, 10 mg/ml of MTT solution was the concentration that gave higher v alues of optical densities with minor medium interference.