Immunofluorescence-based assays have been developed to detect and quantitate Cryptosporidium parvum infection in cell culture. Here, we describe a method that tracks and quantifies the early phase of attachment and invasion of C. parvum sporozoites using a fluorescent dye. Newly excysted sporozoites were labeled with the amine-reactive fluorescein probe carboxyfluorescein diacetate succinimidyl esters (CFSE) using an optimized protocol. The initial invasion of cells by labeled parasites was detected with fluorescent or confocal microscopy. The infection of cells was quantified by flow cytometry. Comparative analysis of infection of cells with CFSE-labeled and unlabeled sporozoites showed that the infectivity of C. parvum was not affected by CFSE labeling. Quantitative analysis showed that C. parvum Iowa and MD isolates were considerably more invasive than Cryptosporidium hominis isolate TU502. Unlike immunofluorescent assays, CFSE labeling permitted the tracking of the initial invasion of C. parvum. Such an assay may be useful for studying the dynamics of host cell–parasite interaction and possibly for drug screening.
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December 2006
THERAPEUTICS-DIAGNOSTICS|
December 01 2006
QUANTITATIVE TRACKING OF CRYPTOSPORIDIUM INFECTION IN CELL CULTURE WITH CFSE
Hanping Feng;
Hanping Feng
Division of Infectious Diseases, Tufts University, Cummings School of Veterinary Medicine, North Grafton, Massachusetts 01536. saul.tzipori@tufts.edu
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Weijia Nie;
Weijia Nie
Division of Infectious Diseases, Tufts University, Cummings School of Veterinary Medicine, North Grafton, Massachusetts 01536. saul.tzipori@tufts.edu
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Ruben Bonilla;
Ruben Bonilla
Division of Infectious Diseases, Tufts University, Cummings School of Veterinary Medicine, North Grafton, Massachusetts 01536. saul.tzipori@tufts.edu
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Giovanni Widmer;
Giovanni Widmer
Division of Infectious Diseases, Tufts University, Cummings School of Veterinary Medicine, North Grafton, Massachusetts 01536. saul.tzipori@tufts.edu
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Abhineet Sheoran;
Abhineet Sheoran
Division of Infectious Diseases, Tufts University, Cummings School of Veterinary Medicine, North Grafton, Massachusetts 01536. saul.tzipori@tufts.edu
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Saul Tzipori
Saul Tzipori
Division of Infectious Diseases, Tufts University, Cummings School of Veterinary Medicine, North Grafton, Massachusetts 01536. saul.tzipori@tufts.edu
* To whom correspondence should be addressed
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J Parasitol (2006) 92 (6): 1350–1354.
Citation
Hanping Feng, Weijia Nie, Ruben Bonilla, Giovanni Widmer, Abhineet Sheoran, Saul Tzipori; QUANTITATIVE TRACKING OF CRYPTOSPORIDIUM INFECTION IN CELL CULTURE WITH CFSE. J Parasitol 1 December 2006; 92 (6): 1350–1354. doi: https://doi.org/10.1645/GE-853R.1
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