Sulforhodamine B (SRB) Assay in Cell Culture to Investigate Cell Proliferation

The SRB assay has been used since its development in 1990 (Skehan et al., 1990) to inexpensively conduct various screening assays to investigate cytotoxicity in cell based studies (Vichai and Kirtikara, 2006). This method relies on the property of SRB, which binds stoichiometrically to proteins under mild acidic conditions and then can be extracted using basic conditions; thus, the amount of bound dye can be used as a proxy for cell mass, which can then be extrapolated to measure cell proliferation. The protocol can be divided into four main steps: preparation of treatment, incubation of cells with treatment of choice, cell fixation and SRB staining, and absorbance measurement. This assay is limited to manual or semiautomatic screening, and can be used in an efficient and sensitive manner to test chemotherapeutic drugs or small molecules in adherent cells. It also has applications in evaluating the effects of gene expression modulation (knockdown, gene expression upregulation), as well as to study the effects of miRNA replacement on cell proliferation (Kasinski et al., 2015).