Preparation of seed organisms for protozoan method development and control studies

Millions of dollars have been spent on detection method development and pilot plant studies for Giardia and Cryptosporidium. Many researchers in this field do not exercise care in the way they prepare and count organisms used as seed in these studies. In many instances seed organisms are purchased and used in experiments without any microscopic observation or determination of suspension density, making data from these studies unreliable and unrepeatable. The method of seed organism preparation, storage, and counting should be optimized for each study design. To be useful in performing repeatable parasite detection methods, accurate enumerations must be done in the range of 10¹ and 10² organisms. Work in this laboratory counting numbers of organisms in this range has shown that the Coulter counter and FACSCalibur methods count with coefficients of variation less than 10% from the mean, while the hemacytometer, chamber slide, and well slide methods count organisms in this range with coefficients of variation up to 80% from the mean.