Neutral red 3 amino 7 dimethylamino 2 methyl phenazine hydrochloride

In vitro cytotoxicity of the stimulatory agents on THP‐1 cells at the concentrations tested was measured using the neutral red uptake assay, following the protocol described by Repetto et al. 68. The neutral red uptake assay is a simple technique used to quantify the proportion of viable cells in culture by addition of the dye neutral red (3‐amino‐7‐dimethylamino‐2‐methyl‐phenazine hydrochloride) (Sigma‐Aldrich), which is incorporated and accumulated in the lysosomes of viable cells. After 24 h of exposure of THP‐1 cells to stimulants, the supernatant was removed and the neutral red assay was performed on the cells. The absorbance of the extracted neutral red from the challenged cells was read in a spectrophotometer at a wavelength at 540 nm, using the blank without cells as the reference.