Adenosine triphosphate (ATP) was measured in whole L3 larvae or pupae (avoiding black pupae) as described in Liu and Lu (2010) (link) Briefly, two larvae or pupae were lysed in 100 μl lysis buffer from ATP Bioluminescence Assay Kit HS II (Merck 11699709001), heated at 95°C for 2 min, and then centrifuged at maximal speed at 4°C for 1 min. 2.5 μl clear supernatant, 187.5 μl dilution buffer, and 10 μl luciferase from the kit were mixed and the luminescence was immediately measured using a Spark Multimode Microplate Reader from Tecan. The absolute ATP amount was calculated according to an ATP standard curve.
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