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Updated: Jun 6, 2026
Development of a Quantitative Recombinase Polymerase Amplification Assay with an Internal Positive Control
Published on: March 30, 2015
Development and evaluation of a real-time PCR assay for quantification of Giardia and Cryptosporidium in sewage
José L Alonso1, Inmaculada Amorós, Irene Cañigral
1Instituto de Ingeniería del Agua y Medio Ambiente, Ciudad Politécnica de la Innovación, Ed. 8G, Acceso D, planta 2, Universidad Politécnica de Valencia, Camino de Vera s/n, 46022 Valencia, Spain. jalonso@ihdr.upv.es
Abstract:
Cryptosporidium and Giardia are major causes of diarrheal disease in humans worldwide and are major causes of protozoan waterborne diseases. Two DNA TaqMan PCR-based Giardia and Cryptosporidium methods targeting a 74-bp sequence of the β-giardin Giardia gene and a 151-bp sequence of the COWP Cryptosporidium gene, respectively, were used as models to compare two different LNA/DNA TaqMan probes to improve the detection limit in a real-time PCR assay. The LNA probes were the most sensitive resulting in 0.96 to 1.57 lower C t values than a DNA Giardia TaqMan probe and 0.56 to 2.21 lower than a DNA Cryptosporidium TaqMan probe. Evaluation of TaqMan Giardia and Cryptosporidium probes with LNA substitutions resulted in real-time PCR curves with an earlier C t values than conventional DNA TaqMan probes. In conclusion, the LNA probes could be useful for more sensitive detection limits.
