Analytical Validation of Quantitative Real-Time PCR Methods for Quantification of Trypanosoma cruzi DNA in Blood Samples from Chagas Disease Patients
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Ramírez, Juan Carlos
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Cura, Carolina Inés
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Cruz Moreira, Otacilio da
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Lages Silva, Eliane
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Juiz, Natalia
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Velázquez, Elsa
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Ramírez, Juan David
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Alberti, Anahi
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Pavia, Paula
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Flores Chávez, María Delmans
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Muñoz Calderón, Arturo
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Pérez Morales, Deyanira
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Santalla, José
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Matta Guedes, Paulo Marcos de
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Peneau, Julie
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Marcet, Paula
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Padilla, Carlos
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Cruz Robles, David
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Valencia, Edward
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Crisante, Gladys Elena
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Greif, Gonzalo
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Zulantay Alfaro, Inés
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Costales, Jaime
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Álvarez Martínez, Miriam
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Martínez, Norma
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Villarroel, Rodrigo
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Villarroel, Sandro
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Sánchez, Zunilda
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Bisio, Margarita
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Parrado, Rudy
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Cunha Galvao, Lucía María da
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Jacome da Camara, Antonia Claudia
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Espinoza, Bertha
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Alarcón de Noya, Belkisyole
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Puerta, Concepción
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Riarte, Adelina
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Diosque, Patricio
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Sosa Estani, Sergio
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Guhl, Felipe
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Ribeiro, Isabela
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Aznar, Christine
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Britto, Constanca
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Yadon, Zaida
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Schijman, Alejandro G.
Admission date
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2015-12-18T12:51:15Z
Available date
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2015-12-18T12:51:15Z
Publication date
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2015
Cita de ítem
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Journal of Molecular Diagnostics Volumen: 17 Número: 5 Páginas: 605-615
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Identifier
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DOI: 10.1016/j.jmoldx.2015.04.010
Identifier
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https://repositorio.uchile.cl/handle/2250/135832
General note
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Artículo de publicación ISI
Sin acceso a texto completo
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Abstract
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An international study was performed by 26 experienced PCR laboratories from 14 countries to assess the performance of duplex quantitative real-time PCR (qPCR) strategies on the basis of TaqMan probes for detection and quantification of parasitic Loads in peripheral blood samples from Chagas disease patients. Two methods were studied: Satellite DNA (SatDNA) qPCR and kinetoplastid DNA (kDNA) qPCR. Both methods included an internal amplification control. Reportable range, analytical sensitivity, limits of detection and quantification, and precision were estimated according to international guidelines. In addition, inclusivity and exclusivity were estimated with DNA from stocks representing the different Trypanosoma cruzi discrete typing units and Tlypanosoma rangeli and Leishmania spp. Both methods were challenged against 156 blood samples provided by the participant laboratories, including samples from acute and chronic patients with varied clinical findings, infected by oral route or vectorial transmission. kDNA qPCR showed better analytical sensitivity than SatDNA qPCR with Limits of detection of 0.23 and 0.70 parasite equivalents/mL, respectively. Analyses of clinical samples revealed a high concordance in terms of sensitivity and parasitic loads determined by both SatDNA and kDNA qPCRs. This effort is a major step toward international validation of qPCR methods for the quantification of T. auzi DNA in human blood samples, aiming to provide an accurate surrogate biomarker for diagnosis and treatment monitoring for patients with Chagas disease.
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Patrocinador
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CONICET
PIP 112-200801-02915
National Agency of Science and Technology grant
PICT 33955