PMID- 25075478
OWN - NLM
STAT- MEDLINE
DCOM- 20141023
LR  - 20190907
IS  - 1678-9849 (Electronic)
IS  - 0037-8682 (Linking)
VI  - 47
IP  - 3
DP  - 2014 May-Jun
TI  - A complete molecular biology assay for hepatitis C virus detection, 
      quantification and genotyping.
PG  - 287-94
LID - S0037-86822014000300287 [pii]
AB  - INTRODUCTION: Molecular biology procedures to detect, genotype and quantify 
      hepatitis C virus (HCV) RNA in clinical samples have been extensively described. 
      Routine commercial methods for each specific purpose (detection, quantification 
      and genotyping) are also available, all of which are typically based on 
      polymerase chain reaction (PCR) targeting the HCV 5' untranslated region (5'UTR). 
      This study was performed to develop and validate a complete serial laboratory 
      assay that combines real-time nested reverse transcription-polymerase chain 
      reaction (RT-PCR) and restriction fragment length polymorphism (RFLP) techniques 
      for the complete molecular analysis of HCV (detection, genotyping and viral load) 
      in clinical samples. METHODS: Published HCV sequences were compared to select 
      specific primers, probe and restriction enzyme sites. An original real-time 
      nested RT-PCR-RFLP assay was then developed and validated to detect, genotype and 
      quantify HCV in plasma samples. RESULTS: The real-time nested RT-PCR data were 
      linear and reproducible for HCV analysis in clinical samples. High correlations 
      (> 0.97) were observed between samples with different viral loads and the 
      corresponding read cycle (Ct - Cycle threshold), and this part of the assay had a 
      wide dynamic range of analysis. Additionally, HCV genotypes 1, 2 and 3 were 
      successfully distinguished using the RFLP method. CONCLUSIONS: A complete serial 
      molecular assay was developed and validated for HCV detection, quantification and 
      genotyping.
FAU - Casanova, Yara Silva
AU  - Casanova YS
AD  - Laboratorio de Diagnostico Molecular, Programa de Pos-Graduacao em Biologia 
      Celular e Molecular Aplicada a Saude, Universidade Luterana do Brasil, Canoas, 
      RS, Brasil.
FAU - Boeira, Thais da Rocha
AU  - Boeira Tda R
AD  - Cachoeirinha, Cachoeirinha, RS, Brasil.
FAU - Sisti, Elisa
AU  - Sisti E
AD  - Laboratorio de Diagnostico Molecular, Programa de Pos-Graduacao em Biologia 
      Celular e Molecular Aplicada a Saude, Universidade Luterana do Brasil, Canoas, 
      RS, Brasil.
FAU - Celmer, Alvaro
AU  - Celmer A
AD  - Cachoeirinha, Cachoeirinha, RS, Brasil.
FAU - Fonseca, Andre Salvador Kazantzi
AU  - Fonseca AS
AD  - Cachoeirinha, Cachoeirinha, RS, Brasil.
FAU - Ikuta, Nilo
AU  - Ikuta N
AD  - Laboratorio de Diagnostico Molecular, Programa de Pos-Graduacao em Biologia 
      Celular e Molecular Aplicada a Saude, Universidade Luterana do Brasil, Canoas, 
      RS, Brasil.
FAU - Simon, Daniel
AU  - Simon D
AD  - Laboratorio de Diagnostico Molecular, Programa de Pos-Graduacao em Biologia 
      Celular e Molecular Aplicada a Saude, Universidade Luterana do Brasil, Canoas, 
      RS, Brasil.
FAU - Lunge, Vagner Ricardo
AU  - Lunge VR
AD  - Laboratorio de Diagnostico Molecular, Programa de Pos-Graduacao em Biologia 
      Celular e Molecular Aplicada a Saude, Universidade Luterana do Brasil, Canoas, 
      RS, Brasil.
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - Brazil
TA  - Rev Soc Bras Med Trop
JT  - Revista da Sociedade Brasileira de Medicina Tropical
JID - 7507456
RN  - 0 (5' Untranslated Regions)
RN  - 0 (DNA Primers)
RN  - 0 (RNA, Viral)
SB  - IM
MH  - 5' Untranslated Regions/*genetics
MH  - DNA Primers
MH  - Genotype
MH  - Hepacivirus/*genetics/isolation & purification
MH  - Hepatitis C/*diagnosis
MH  - Humans
MH  - Polymorphism, Restriction Fragment Length
MH  - RNA, Viral/*blood
MH  - Reverse Transcriptase Polymerase Chain Reaction
MH  - Sensitivity and Specificity
MH  - Viral Load
EDAT- 2014/07/31 06:00
MHDA- 2014/10/24 06:00
CRDT- 2014/07/31 06:00
PHST- 2014/02/26 00:00 [received]
PHST- 2014/06/16 00:00 [accepted]
PHST- 2014/07/31 06:00 [entrez]
PHST- 2014/07/31 06:00 [pubmed]
PHST- 2014/10/24 06:00 [medline]
AID - S0037-86822014000300287 [pii]
AID - 10.1590/0037-8682-0040-2014 [doi]
PST - ppublish
SO  - Rev Soc Bras Med Trop. 2014 May-Jun;47(3):287-94. doi: 
      10.1590/0037-8682-0040-2014.