Cytotoxicity of FCN has been studied extensively and some of the results are shown in Figure 3.8. Cytotoxicity study has been performed using methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay and Trypan blue assay. Typically, cells are exposed to FCN for 24 h in the concentration range of 0.1–1 mg/mL, which is ~100- to 1000-times higher than required for imaging application. Next, MTT solution was added to each well 4 h before the end of the incubation. The medium was discarded and the produced formazan was dissolved with DMSO. The plates were read with absorbance at 550 nm.This reduction reaction only occurs in live cells, and so corresponds to a measure of cell viability. The cells are treated with isopropanol organic solvent to extract formazan, which is measured spectrophotometrically at 490 nm.