Mouse antirabbit igg

Ecdysteroid titers were determined by a 20E EIA kit (Cayman Chemical) according to the manufacturer’s protocol. At 24 h after infectious blood meal, individual female mosquitoes were stored in 100 µl of 100% methanol at −80°C. The mosquito whole body was homogenized and kept at 4°C for 30 min before centrifugation (10,000 ×g, 10 min). The supernatant was transferred to a new 1.5-ml Eppendorf tube, and the solvent was removed by centrifugation at reduced pressure. The extracted total ecdysteroids were redissolved in 50 µl of EIA buffer, and loaded on the 96-well strip plate precoated with mouse anti-rabbit IgG (Cayman Chemical). Samples in duplicate were incubated with 50 µl tracer and 50 µl 20E antiserum overnight at 4°C and then developed with 200 µl Ellman’s Reagent for 90–120 min. Absorbance was measured by a BioTek Synergy Mx microplate reader at 412 nm.