OPTIMIZATION OF THE SULFORHODAMINE-B COLORIMETRIC ASSAY

Sulforhodamine B (SRB) protein staining has been widely used for cell proliferation and chemosensitivity testing, substituting for tetrazoli um-based assays. However, the cell fixation step in the original assay is subject to error. We tested whether aspiration of medium with an a utomatic microplate multiwash device prior to fixation improves the me thod for adherent cells. A panel of adherent cell lines was used. Sign al-to-noise ratios were significantly increased in the new assay. Coef ficients of variation (CV) between replicate wells were significantly lower especially at lower cell densities. The linearity of the method improved, with absolute linearity over the whole range of cell densiti es. The aspiration procedure dislodged only negligible numbers of cell s. Cytotoxicity testing using the cytotoxic agent paclitaxel showed no IC50 (50% inhibitory concentration) differences between the new and o riginal methods but a better CV was associated with the optimized prot ocol. We conclude that aspiration of the growth medium prior to fixing comprises a safe and reliable practice which improves CV, linearity a nd the signal-to-noise ratio of the SRB assay. (C) 1997 Elsevier Scien ce B.V.