Detection of pyroptosis by measuring released lactate dehydrogenase activity

Pyroptosis is a form of programmed, inflammatory cell death that is dependent on the activation of a cysteine protease caspase-1. Following caspase-1 activation via inflammasomes (including NLRP3, NLRC4, Nlrp1b, and AIM2), cells lose membrane integrity and lyse, releasing lactate dehydrogenase (LDH) that is normally maintained within the cell cytosol. Thus, pyroptosis is distinct from apoptosis, which results in cellular contents being enclosed within membrane blebs during cellular demise. LDH is only released from apoptotic blebs after secondary necrosis occurs. Pyroptosis is distinct from necrosis in that it requires the activity of caspase-1. In this chapter, we describe enzymatic assays for the detection of LDH released by pyroptotic cells using a commercially available kit, as well as a simple and cost-effective method adapted from Decker et al. (J Immunol Methods 115:61-69, 1988).