Active substances: Doxycycline
For direct blood feeding on birds, the downy breast feathers were trimmed, and the exposed bird breast was placed on top of the mosquito cage.
The birds were gently restrained for 30 minutes while the mosquitoes blood fed through the mosquito cage organdy. Given the difficulties of direct mosquito blood feeding on live chickens, supplemental serum-replacement membrane blood feeds were also performed, where frozen chicken serum was used in reconstituted blood meals using red blood cells from defibrinated calf blood.
IVM was extracted from serum following methanol precipitation. Samples were centrifuged for 30 min at 16,000 x g. Supernatants were transferred and evaporated to dryness using a Speedvac concentrator Savant.
Samples were derivatized according to previously published literature. Excitation and emission spectra were 365 and 470 nm, respectively.
Instrument CV was 6. Intraday CV ranged between 4. Interday reproducibility was 15.
Retention time CV was 1. The method was linear across the range of concentrations tested in the standard curve 3. Linear regression curves containing fortified IVM serum samples with concentrations of 3.
Limits of detection and quantification were 1.