| 1. Ensure procedures are in place to maintain samples at 37°C at all times from venipuncture to the end of the membrane feeding experiment, in order to prevent activation of gametocytes and reduction of infectivity. |
| 2. Use commercially available membranes that are more amenable to standardization than animal skin. |
| 3. Set an appropriate the time range that mosquitoes are allowed to feed on a blood meal. This is a purely ethical issue in direct skin feeding assays but may affect the validity of membrane feeding assays since the infectiousness of gametocytes may decline during membrane feeding experiments. A feeding time of 15 minutes is commonly used. |
| 4. Keep mosquito number/cup size/age of mosquitoes and also mosquito husbandry in feeding experiments as constant as possible. |
| 5. Remove unfed mosquitoes after the feeding experiment (rather than selecting fed mosquitoes). |
| 6. Estimate the assay variability by repeating the same experiment multiple times and incorporate this information in power calculations and statistical analysis. |
| 7. Track the number of dead mosquitoes prior to examination to determine differences in mosquito survival between cages/cups or between gametocyte donors. |
| 8. Maximize the number of mosquitoes that can be used in experiments to ensure enough are dissected for achieving sufficient statistical power to address research questions. |
