Toxglo assay kit

Following MZ treatment, nematodes were washed as described. Worms were then counted to yield 10 live nematodes/μL per treatment group, and 200 μL (200 worms) was added to each well. Per the protocol provided by Promega, relative ATP concentration was measured using the Promega Mitochondrial ToxGlo™ Assay (Promega Corporation, Madison, WI). Initially, 20 μL of a fluorogenic peptide substrate, provided in the ToxGlo™ assay kit (Promega Corporation, Madison, WI) was added, and incubated at 20°C for 60 min to determine percent live worms compared to control. Fluorescence was measured at an excitation wavelength of 495 nm_Ex and an emission wavelength of 520–530 nm_Em using a Promega^® GloMax-Multi⁺ Detection System. Next, 100 μL of a luciferin-containing ATP detection reagent, also provided in the ToxGlo™ assay kit (Promega Corporation, Madison, WI), was added to each well and incubated at room temperature for 15 min. Luminescence was measured using the Promega^® GloMax-Multi⁺ Detection System.