In-house standardization and validation of a multiplex RT-PCR assay for the detection of 13 respiratory viruses

Hernán Vargas; Ángela Diaz; Yamile Celis; Liliana Díaz; Sandra Gómez; Jenny Sánchez; Carlos Golijow; Patricia Arce

doi:10.22490/24629448.1746

Vol. 14 Núm. 26 (2016)

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2016-12-15

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In-house standardization and validation of a multiplex RT-PCR assay for the detection of 13 respiratory viruses

DOI: https://doi.org/10.22490/24629448.1746

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Hernán Vargas Secretaría Distrital de Salud; Bogotá, Colombia.

Ángela Diaz Secretaría Distrital de Salud; Bogotá, Colombia.

Yamile Celis Secretaría Distrital de Salud; Bogotá, Colombia.

Liliana Díaz Secretaría Distrital de Salud; Bogotá, Colombia.

Sandra Gómez Secretaría Distrital de Salud; Bogotá, Colombia.

Jenny Sánchez Secretaría Distrital de Salud; Bogotá, Colombia.

Carlos Golijow

Patricia Arce Secretaría Distrital de Salud; Bogotá, Colombia.

Background. Multiplex real time PCR is increasingly used to diagnose respiratory viruses and has shown to be superior to traditional methods, such as culture and antigen detection. Objective. Standardization and validation of a multiplex real-time PCR assay for the detection of 13 respiratory viruses. Methods. The assay was validated using RNA control targets and comparing results to single-target PCR’s. Results. Using RNA controls the multiplex format was found to be as sensitive and specific as the single-target PCRs, and no competition was observed between targets. The efficiencies for most of the reactions were approximately 90%, but a lower efficiency was found for Parainfluenza 2 with a rate of amplification in each cycle of 86.63%. On the other hand, a higher efficiency was observed in respiratory syncytial virus A and respiratory syncytial virus B ((93.07% each). Conclusion: This multiplex RT-PCR format shows an adequate efficiency, demonstrating an excellent sensitivity, specificity and repeatability for all the studied respiratory viruses.

Multiplex Real Time PCR, Respiratory virus, Standardization.

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Vargas, H., Diaz, Ángela, Celis, Y., Díaz, L., Gómez, S., Sánchez, J., Golijow, C., & Arce, P. (2016). In-house standardization and validation of a multiplex RT-PCR assay for the detection of 13 respiratory viruses. Nova, 14(26), 9-15. https://doi.org/10.22490/24629448.1746

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