Ready-to-use qPCR for detection of
| Autor: | Alexandre D T, Costa, Thiago, Jacomasso, Elaine C, Mattos, Aline B, Farias, Rita C P, Rampazzo, Rebeka S, Pinto, Walleyd, Tassi, Maria Aparecida M, Marciano, Vera Lucia, Pereira-Chioccola, Helen R, Murphy, Alexandre J, da Silva, Marco A, Krieger |
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| Rok vydání: | 2020 |
| Předmět: | |
| Zdroj: | Food and Waterborne Parasitology |
| ISSN: | 2405-6766 |
| Popis: | Foodborne outbreaks caused by parasites have long been a public health issue. Among the available contamination detection methods, qPCR is one of the most sensitive and specific. However, it can be cumbersome and error-prone, if used by unexperienced users. Moreover, qPCR reagents usually require freezer temperatures for transportation and storage. We present a gelified reaction format that allows the reagents to be stored at 2–8 °C for up to 90 days without losing performance. The gelification process eliminates most operator mistakes during reaction setup, and renders the qPCR plates ready-to-use. The new reaction makeup was evaluated using artificially contaminated samples of distinct food matrices for sensitivity, specificity, repeatability, reproducibility, and stability. Samples consisted of cilantro leaves and raspberry fruits spiked with Cyclospora cayetanensis oocysts, as well as açai pulp and sugarcane juice tainted with Trypanosoma cruzi trypomastigotes. No significant difference between the gelified and the non-gelified qPCR was found. Our results suggest that gelifying the assay may help to achieve more reproducible qPCR data across laboratories, thus supporting surveillance actions. (170 words) Highlights • The ready-to-use qPCR has the same performance as the freezer-stored reagents. • The ready-to-use qPCR set up is faster and the overall process is up to 20% cheaper. • Proposed endogenous reaction control target is ubiquitous in higher plants. |
| Databáze: | OpenAIRE |
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