4.2.2 (b) Lactate Dehydrogenase (LDH) release assay to identify cell cytotoxicity
Lactate dehydrogenase is a cytosolic enzyme which is released into the cell culture media with plasma membrane damage which can be measured. Levels of LDH released in the media are an indicator whether the cells are undergoing necrosis. High levels of LDH in the media are proportional to cell death via necrosis. Plates with unstimulated cells and Rv2615c or control (LPS/Staurosporine) stimulated cells were centrifuged and 20µl of media from each well and 20µl reaction buffer were added to fresh wells. Plates were incubated for 30 min at room temperature following addition of 20µl stop solution. Absorbance reading was recorded at 490nm.