Production of anti-Cryptosporidium polyclonal antibodies and standardization of direct immunofluorescence for detecting oocysts in water

• Published in: Revista da Sociedade Brasileira de Medicina Tropical
• Main Authors: Silvia Cristina Osaki, Adriana Oliveira Costa, Ludmilla Della Coletta Troiano, Ernesto Renato Kruger, Juliana Tracz Pereira, Nelson Luis Mello Fernandes, Márcia Benedita de Oliveira Silva, Vanete Thomaz Soccol
• Format: Article in Journal/Newspaper
• Language: English
• Published: Sociedade Brasileira de Medicina Tropical (SBMT) 2011
• Subjects: Anticorpos policlonais; Imunofluorescência direta; Oocistos; Água; Arctic medicine. Tropical medicine; RC955-962; Arctic
• Online Access: https://doi.org/10.1590/S0037-86822011000500011; https://doaj.org/article/9ac2a2077e6b4b5b845a5c8873617e8a

INTRODUCTION: The production of anti-Cryptosporidium polyclonal antibodies and its use in direct immunofluorescence assays to determine the presence of Cryptosporidium in water are described in the present work. METHODS: Two rabbits were immunized with soluble and particulate antigens from purified Cryptosporidium oocysts. The sera produced were prepared for immunoglobulin G extraction, which were then purified and conjugated with fluorescein isothiocyanate (FITC). Slides containing known amounts of oocysts were prepared to determine the sensitivity of the technique. To test the specificity, slides containing Giardia duodenalis cysts were prepared. RESULTS: The conjugate was successfully used in water samples experimentally contaminated with Cryptosporidium oocysts, and it was possible to detect up to five oocysts/spot, corresponding to contamination of 250 oocysts/mL. CONCLUSIONS: The three immunizations performed in the rabbits were enough to produce antibodies against Cryptosporidium, the standard direct immunofluorescence assay permitted the detection of five oocysts in 20% of the samples, and no cross-reaction with Giardia duodenalis cysts occurred.