title: Evaluation of an alternative IMS dissociation procedure for use with Method 1622: detection of Cryptosporidium in water.
abstract: 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 4',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 degrees 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.
authors: National Exposure Research Laboratory, U.S. EPA, 26 W. Martin Luther King Drive, Cincinnati, OH 45268-1320, USA. ware.michael@epa.gov
authors: Michael W
authors: MW
authors: Ware
authors: Larry
authors: L
authors: Wymer
authors: H D Alan
authors: HD
authors: Lindquist
authors: Frank W
authors: FW
authors: Schaefer
mesh_terms: D000818:Animals; D016785:Cryptosporidium parvum / Q000302:isolation & purification*; D004784:Environmental Monitoring / Q000379:methods*; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique / Q000379:methods; D005456:Fluorescent Dyes / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D018189:Immunomagnetic Separation / Q000379:methods*; D007211:Indoles / Q000378:metabolism; D015999:Multivariate Analysis; D033661:Oocysts / Q000302:isolation & purification; D014481:United States; D014484:United States Environmental Protection Agency; D014867:Water / Q000469:parasitology*
chemical_list: D005456:Fluorescent Dyes; D007211:Indoles; D014867:Water; C007293:DAPI