The objective of this study was to utilize defined protocols to evaluate three different fluorogenic vital dyes assays (4', 6' diamidino-2-phenyl indole/propidium iodide, SYTO-9 and SYTO-59) with a maximized excystation procedure and neonatal mouse infectivity. The laboratories participating in this study were all experienced in either viability and/or infectivity assays; nonetheless, individuals from these laboratories were cross-trained and standard operating protocols were developed and distributed to ensure standardized methodologies for viability and infectivity assays in both the US and United Kindgom. Utilizing these protocols, the same isolate of fresh oocysts (<1 month) was used to develop standard curves for mouse infectivity assays in both the US and United Kingdom. This was followed by partial-inactivation experiments with fresh oocysts, using pulsed field ultraviolet light and ozone, in both the US and United Kingdom. Oocyst inactivation levels were measured using the three vital dye assays, maximized in vitro excystation and neonatal mouse infectivity assays.