Immunofluorescent Staining of Giardia and Cryptosporidium on Cellulose Acetate Membranes: A Major Source of Variability in the ICR Protozoan Method

摘要

Three studies were conducted to investigate the reason for the loss of countablernorganisms in the Information Collection Rule (ICR) protozoan method. In the first study,rn35 laboratories representing 61 principal analysts (PAs) who participated in an initialrnround of laboratory approvals, and who had completed their first set of protozoanrnperformance evaluation (PE) samples, received QC vials containing known concentrationsrnof Giardia cysts and Cryptosporidium oocysts. Analysts were instructed to stain andrncount 3 membranes using the ICR method. The recovery of cysts and oocysts wasrncalculated and these data were compared to their PE approval results. After analysts whornhad failed the first round had the opportunity to analyze a repeat set of PE samples, therndata were re-evaluated to determine if there was a correlation between membrane filterrnrecoveries and the second round PE results. The average recovery of Giardia cysts andrnCryptosporidium oocysts seeded directly on membranes was 69% ± 20% and 42% ±rn22%, for cysts and oocysts, respectively. If PE results were based solely on thernacceptable recovery of Giardia cysts, 46 of the 61 PA’s (75%) in this survey would havernpassed their PE samples. In addition, 40 of those who would have passed, recoveredrnmore than 61% of the cysts seeded directly on sample membranes, suggesting arncorrelation between membrane recoveries and PE sample performance. Recoveries ofrnoocysts on sample membranes and PE sample performance were poorly correlated. Thernrange of membrane recoveries for Cryptosporidium was 11% to 80% for the 39 PA’s whornpassed their PE samples after 1 or 2 attempts.