A dot-enzyme-linked immunosorbent assay, dot-ELISA, that allows for identification of human blood meals in mosquitoes in less than 2 h is presented. Strips of mylar-backed nitrocellulose paper, the dipstick, may be inoculated with anti-human capture antibody, blocked, dried and stored for at least 3 months before use. The Dipstick ELISA consistently detected human blood meals at 24 h post-blood meal in frozen Anopheles mosquitoes and at 32 h post-blood meal on samples eluted off filter-paper smears. This ELISA detects human IgG at dilutions of 1:25,600, and produces strong positive results at dilutions between 1:400 and 1:12,800. The Dipstick ELISA is highly specific, and no false positives were detected when tested against cow, horse, goat, dog, cat, pig, rabbit, mouse, rat, chicken, raccoon and opossum sera. All reagents for the assay are commercially available. The Dipstick ELISA meets requirements for a rapid and simple assay for the identification of human blood meals and should have particular application to short-term field studies and emergency epidemiological investigations. A modified protocol, the Trough ELISA, which treats dipsticks jointly in disposable pipet troughs during the conjugate and substrate steps, was developed and produced as a kit. The Trough ELISA produces 1-4% random false positives, and we recommend that the Trough ELISA not be used.
