The accurate determination of the presence of Giardia cysts and Cryptosporidium oocysts in surface waters requires a reliable method for the detection and enumeration of these pathogenic organisms. Published methods have usually reported recovery efficiencies of less than 50% for both cysts and oocysts. Typically, the losses are greater for Cryptosporidium oocysts than they are for Giardia cysts. The purpose of this study was to examine procedures used for sample collection, elution, concentration, and clarification to determine when losses of cysts and oocysts occurred during processing. The results showed that major losses of cysts and oocysts occurred during centrifugation and clarification. Depending on the centrifugation force, oocyst losses of as high as 30% occurred for each centrifugation step. A 1.15-specific-gravity Percoll-sucrose gradient was needed to optimize recovery of oocysts from natural water samples. Minor improvements in the procedure could be accomplished by selecting a filter other than the recommended 1-m-pore-size (nominal-porosity) polypropylene filter. Giardia cysts and Cryptosporidium oocysts are environmentally resistant intestinal parasites that can cause gastroenteritis in humans when they are ingested. These organisms can be transported by water and have caused documented waterborne outbreaks of giardiasis and cryptosporidiosis (6). The method of detection for Giardia cysts and Cryptosporidium oocysts in water samples relies primarily on microscopic observation of water samples by an immunofluorescence technique (1). This method involves filtration of a large volume of water through a 1-m-pore-size (nominal-porosity) cartridge filter, elution of the microorganisms from the filter using a detergent-based elution medium, concentration by centrifugation, clarification using a 1.10-specific-gravity Percoll-sucrose density gradient, indirect fluorescent-antibody labeling, and microscopic examination of the sample. The immunofluorescence assay (IFA) for Giardia cysts and Cryptosporidium oocysts is time-consuming and labor intensive and requires a large degree of analytical expertise. Moreover, the recovery efficiency of the procedure, especially for Cryptosporidium oocysts, is relatively low. LeChevallier et al. (5) reported an average method efficiency of 68.6% for Giardia cysts in seeded tap water; however, Cryptosporidium oocysts were recovered at an average level of only 25.3%. A recent study of 12 commercial laboratories that processed spiked proficiency samples showed an average of 9% recovery of Giardia cysts and 3% recovery of Cryptosporidium oocysts (2). Moreover, 36% failed to detect Giardia cysts and 55% failed to find Cryptosporidium oocysts, even though the samples contained 99 cysts and 80 oocysts. Large variations in recovery efficiencies have been reported between different laboratories and even within a single laboratory (2, 3). The objective of the present study was to analyze the IFA technique, to determine when the losses of cysts and oocysts occurred during sample pr...