Measuring Cell Death Using LDH Assay

LDH release from the cell was used as an indicator of cell death using an NAD⁺ reduction assay (Roche Applied Science). Supernatants from treated cells were collected, clarified by centrifugation at 400 g for 5 min, filtered and used for the assay. For a positive control, total LDH content in untreated monocytes was obtained by lysing cells with 1% Triton X-100. RPMI-1640 media was used as a blank and OD values were subtracted from readings of samples and positive control. LDH concentration in the medium was measured at 490 nm. Cell death was calculated by the formula: cytotoxicity (%) = [(sample-blank)/(positive control-blank) × 100], as described earlier (Gavrilin et al., 2012 (link)).