Novel Methodology for the Detection of Cryptosporidium parvum: A Comparison of Cooled Charge Couple Devices (CCD) and Flow Cytometry

Abstract

Flow cytometry and CCD were assessed for their usefialness in the detection of oocysts of Cryptosporidium. Oocysts were labelled with FITC-monoclonal antibody and with the nuclear stains 4’6-diamidino-2-phenyl indole (DAPI) and propidium iodide (PI) before analysis by flow cytometer and CCD. Although the flow cytometer tested was able to concentrate particles and place them on a slide for subsequent viewing, readily sorting oocysts from contaminating debris, confirmation by fluoresence microscopy was still essential, even when additional parameters such as the inclusion of DAPI is used. Initial observations from the use of CCD, however, suggested that screening samples for oocysts was a possibility. Three dimensional visualisation of individual oocysts can map precisiely both the detailed morphology and the exact size of oocysts, thereby making confirmation by fluoresence microscopy unneccesary.