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A flow cytometric protocol for detection of Cryptosporidium spp.
Author(s) -
Barbosa Joana M. M.,
CostadeOliveira Sofia,
Rodrigues Acácio G.,
Hanscheid Thomas,
Shapiro Howard,
PinaVaz Cidália
Publication year - 2008
Publication title -
cytometry part a
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.316
H-Index - 90
eISSN - 1552-4930
pISSN - 1552-4922
DOI - 10.1002/cyto.a.20502
Subject(s) - cryptosporidium , cryptosporidium parvum , monoclonal antibody , flow cytometry , detection limit , biology , staining , microbiology and biotechnology , antibody , apicomplexa , protozoa , chromatography , chemistry , protozoal disease , immunology , genetics , malaria , feces
Cryptosporidium parvum is transmitted through water and can cause severe diarrhea. The diagnosis is usually based upon observer‐dependent microscopic detection of oocysts, with rather low sensitivity and specificity. Our objective was to optimize a flow cytometric (FC) protocol for the detection of C. parvum . A specific monoclonal antibody conjugated with R‐phycoerythrin was incubated with dead oocysts to determine the optimal antibody concentration. Serial concentrations of oocysts were stained with the optimized concentration and analyzed by FC. The lower detection limit was determined, and the possibility of cross‐reaction was investigated using prokaryotic and eukaryotic microorganisms. A FC protocol was optimized to detect oocysts in spiked human stools. The optimal antibody concentration was found to be 3.0 μg/ml. The lowest number detectable was 2 × 10 3 oocysts/ml. Staining procedure was specific, as no cross‐reactions were observed. This reliable and easy FC protocol allow the specific detection of Cryptosporidium oocysts, even at very low concentrations, which is important for public health and further studies of treatment efficacy. © 2007 International Society for Analytical Cytology.