U.S. Environmental Protection Agency


Date of this Version



Published in Journal of Microbiological Methods 55 (2003) 575– 583. DOI:10.1016/j.mimet.2003.06.001


U.S.EPA Methods 1622 and 1623 are used to detect and quantify Cryptosporidium oocysts in water. The protocol consists of filtration, immunomagnetic separation (IMS), staining with a fluorescent antibody, and microscopic analysis. Microscopic analysis includes detection by fluorescent antibody and confirmation by the demonstration of 1–4 sporozoites or nuclei after staining with 4V,6-diamidino-2-phenyl indole dihydrochloride (DAPI). The purpose of this study was to evaluate a new IMS dissociation, a 10-min incubation at 80 °C. Heat dissociation improved the average oocyst recovery from 41% to 71% in seeded reagent water, and from 10% to 51% in seeded river samples. The average DAPI confirmation rate improved from 49% to 93% in reagent water, and from 48% to 73% in river samples. This modification improved both oocyst recovery and confirmation.