Fatty acid methyl esters (FAMEs) were produced from housefly (Musca domestica L.) larvae fed by poultry manure. The larval lipid output was 23.6 g from 1 kg poultry (107.3 g fresh larva), under conditions: inoculum density of 8000 larvae/kg, 28 degrees C, and 4 days. A practical method involving physical refining were developed specifically for housefly larval lipid (acid value = 15.1 mg KOH/g). With evaporator temperature of 155 degrees C, fatty acids distilled from molecular distillation were esterified with methanol catalyzed by benzenesulfonic acid (BSA) and residue, acylglycerols (fatty acids removal rate: 96.96% wt.), were transesterified with methanol using NaOH catalyst to produce fatty acid methyl ester biodiesel. Residual alkali catalyst in glycerol after transesterification was used to neutralize the acid catalyst and FFA from esterification. Acid value (AV) of esterification product (2.72 mg KOH/g) were lowered to 0.34 mg KOH/g. Biodiesel produced from housefly larval oil (HLO) satisfies the ASTM D6751-10 standard.