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

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. Two rabbits were immunized with soluble and particulate antigens from purified Cryptosporidium oocysts...

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Bibliographic Details
Published in:Revista da Sociedade Brasileira de Medicina Tropical 2011-10, Vol.44 (5), p.587-590
Main Authors: Osaki, Silvia Cristina, Costa, Adriana Oliveira, Troiano, Ludmilla Della Coletta, Kruger, Ernesto Renato, Pereira, Juliana Tracz, Fernandes, Nelson Luis Mello, Silva, Márcia Benedita de Oliveira, Soccol, Vanete Thomaz
Format: Article
Language:English
Subjects:
Animals
Antibodies, Protozoan - biosynthesis
Anticorpos policlonais
Cryptosporidium
Cryptosporidium - immunology
Cryptosporidium - isolation & purification
Fluorescent Antibody Technique, Direct - standards
Fresh Water - parasitology
Immunization
Imunofluorescência direta
Oocistos
Oocysts
Oocysts - immunology
Rabbits
Sensitivity and Specificity
TROPICAL MEDICINE
Water analysis
Water pollution
Water sampling
Água
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Summary: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. 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. 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. 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.
ISSN:0037-8682
1678-9849
1678-9849
0037-8682
DOI:10.1590/S0037-86822011000500011