Open AccessAn immunomagnetic separation polymerase chain reaction assay for rapid and ultra‐sensitive detection of Cryptosporidium parvum in drinking water
Author(s) -
HallierSoulier Sylvie,
Guillot Emmanuelle
Publication year - 1999
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.1999.tb13674.x
Subject(s) - immunomagnetic separation , polymerase chain reaction , chromatography , cryptosporidium parvum , oligomer restriction , chemistry , digoxigenin , microbiology and biotechnology , oligonucleotide , biology , dna , biochemistry , in situ hybridization , gene , gene expression
A sensitive and rapid method was developed to detect Cryptosporidium parvum oocysts in drinking water. This molecular assay combined immunomagnetic separation with polymerase chain reaction amplification to detect very low levels of C. parvum oocysts. Magnetic beads coated with anti‐cryptosporidium were used to capture oocysts directly from drinking water membrane filter concentrates, at the same time removing polymerase chain reaction inhibitory substances. The DNA was then extracted by the freeze‐boil Chelex‐100 treatment, followed by polymerase chain reaction. The immunomagnetic separation‐polymerase chain reaction product was identified by non‐radioactive hybridization using an internal oligonucleotide probe labelled with digoxigenin. This immunomagnetic separation‐polymerase chain reaction assay can detect the presence of a single seeded oocyst in 5–100‐l samples of drinking water, thereby assuring the absence of C. parvum contamination in the sample under analysis.