title: The use of real-time PCR to detect hepatitis C virus RNA in dried blood spots from Brazilian patients infected chronically.
abstract: Collecting and transporting samples for RNA analysis can be challenging, especially in situations where financial resources are limited. In this study, a quantitative real-time PCR (qPCR) for the analysis of HCV RNA was developed and adapted for use with dried blood spot (DBS) samples. A qPCR for HCV 5'NCR, an internal control and a calibration curve were developed, and the sensitivity, specificity and dynamic range of amplification were evaluated using a panel of viruses. Plasma and DBS samples from 100 patients who had completed four weeks of Peginterferon alfa-2b+Ribavirin treatment were collected (DBS on SS903 collection cards and transported at room temperature). After 24 weeks of treatment, samples were collected from 68 of these patients. Of the 168 samples, 2 yielded false-negative results, and 4 yielded false-positive results (sensitivity was 98%, specificity was 94.3%, positive predictive value was 96.1%, and negative predictive value was 96.9%). Additionally, 2039 DBS samples from 1114 patients currently undergoing treatment for a chronic HCV infection in a clinical trial were tested. Only 10 samples out of the 2039 yielded invalid results warranting re-collection of DBS. The detection of HCV RNA in DBS can be a cost-effective strategy for HCV treatment monitoring, especially in settings where resources are limited.
authors.affiliation: Laboratorio Centro de Genomas, Sao Paulo, Brazil.
authors.forename: Carlos
authors.initials: C
authors.lastname: Santos
authors.forename: Alexanda
authors.initials: A
authors.lastname: Reis
authors.forename: Cintia Vilhena
authors.initials: CV
authors.lastname: Dos Santos
authors.forename: Cristine
authors.initials: C
authors.lastname: Damas
authors.forename: Mariliza Henrique
authors.initials: MH
authors.lastname: Silva
authors.forename: Mônica Valverde
authors.initials: MV
authors.lastname: Viana
authors.forename: Maria Lucia
authors.initials: ML
authors.lastname: Ferraz
authors.forename: Dimas
authors.initials: D
authors.lastname: Carnauba
authors.forename: Fabiane
authors.initials: F
authors.lastname: El-Far
authors.forename: Fernando
authors.initials: F
authors.lastname: Serra
authors.forename: Ricardo Sobhie
authors.initials: RS
authors.lastname: Diaz
mesh_terms: D000998:Antiviral Agents / Q000008:administration & dosage; D001769:Blood / Q000821:virology*; D001938:Brazil; D003890:Desiccation; D003951:Diagnostic Errors; D016903:Drug Monitoring / Q000379:methods; D016174:Hepacivirus / Q000235:genetics / Q000302:isolation & purification*; D019698:Hepatitis C, Chronic / Q000175:diagnosis* / Q000188:drug therapy / Q000821:virology; D006801:Humans; D000077190:Interferon alpha-2; D016898:Interferon-alpha / Q000008:administration & dosage; D025202:Molecular Diagnostic Techniques / Q000379:methods; D011092:Polyethylene Glycols / Q000008:administration & dosage; D012367:RNA, Viral / Q000235:genetics / Q000302:isolation & purification*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D011994:Recombinant Proteins / Q000008:administration & dosage; D012254:Ribavirin / Q000008:administration & dosage; D012680:Sensitivity and Specificity; D013048:Specimen Handling / Q000379:methods*; D014773:Virology / Q000379:methods
chemical_list: D000998:Antiviral Agents; D000077190:Interferon alpha-2; D016898:Interferon-alpha; D012367:RNA, Viral; D011994:Recombinant Proteins; D011092:Polyethylene Glycols; D012254:Ribavirin; C417083:peginterferon alfa-2b