Photometric recording of cell viability using trypan blue in perfused cell cultures

A perfusion system was developed to increase the reliability of cell viability estimations by continuous measurement of the uptake of trypan blue dye. Monolayer cell cultures were perfused with buffer containing toxic substances and trypan blue, and the staining of cells was continuously recorded at 591 nm in a spectrophotometer. Using mercuric chloride and methylmercuric chloride as test substances with C6 rat glioma cells, time- and dose-dependent increases in light absorbance were obtained over a 12h recording period. Methylmercuric chloride at 10(-6) M caused a half-maximal increase in relative absorbance in 4.5 h, whereas the corresponding time for mercuric chloride was 10.5 h.