Measurement of LDH and Caspase 3/7 in RSV-Infected Cells

LDH and caspase 3/7 were measured in the supernatants of RSV-infected HEp-2 and A549 cells as previously described (20 (link)). In brief, total LDH activity was measured in the supernatant using the LDH Cytotoxic Detection Kit Plus, (Roche Applied Science, Indianapolis, IN, USA) following protocol instructions. l-lactate dehydrogenase (Roche Applied Science) was used to construct a standard curve that demonstrated linear dynamic range (r = 0.998) at the dilutions tested from 3.9 to 125 milliunits per milliliter (mU/mL). Caspase 3/7, a marker of apoptosis, was measured using the Caspase-Glo-3/7 kit (Promega, Madison, WI, USA). The luminescence assay was measured using a Biotek Synergy H1 microplate reader (Biotek) and reported as relative luminescence units (RLU). Purified caspase 3 from ENZO (catalog no. BML-SE169) was used as the standard for Caspase Glo 3/7 assays and Gen5 Imager Software was used for analysis.