title: Development of a low-cost real-time reverse transcriptase-polymerase chain reaction technique for the detection and quantification of hepatitis C viral load.
abstract: BACKGROUND
  It is necessary to develop a highly specific and sensitive assay to quantify the exact amount of hepatitis C virus (HCV) RNA in blood of patients with hepatitis C. For this reason, a real-time reverse transcriptase-polymerase chain reaction (RT-PCR) assay for quantification of HCV RNA in human plasma was developed.
  
  
  METHODS
  A pair of primers as well as hybridization probes were selected. A real-time RT-PCR was set up and optimized. To establish the sensitivity of the assay, a serial dilution of HCV standards and reference sera, including the six major HCV genotypes, was used. The performance of the assay was evaluated with 191 known HCV-RNA positive and 100 negative samples.
  
  
  RESULTS
  The real-time assay had a sensitivity of 50 IU/mL, with a dynamic range of detection between 10(3) and 10(6) IU/mL. The coefficients of variation of threshold cycle values in intra- and inter-day-runs were <1.77% and 3.40%, respectively. Measurement of HCV-RNA positive samples yielded reproducible data with 100% specificity.
  
  
  CONCLUSIONS
  The high sensitivity, simplicity, reproducibility, wide dynamic range, and low cost of this real-time HCV RNA quantification makes this method especially suitable for monitoring viral load during therapy and tailoring of treatment schedules accordingly.
authors.affiliation: Digestive Disease Research Center, Shariati Hospital, Tehran University of Medical Sciences, Tehran, Iran.
authors.forename: Kiana
authors.initials: K
authors.lastname: Shahzamani
authors.forename: Shahin
authors.initials: S
authors.lastname: Merat
authors.forename: Houri
authors.initials: H
authors.lastname: Rezvan
authors.forename: Siamak
authors.initials: S
authors.lastname: Mirab-Samiee
authors.forename: Hooman
authors.initials: H
authors.lastname: Khademi
authors.forename: Reza
authors.initials: R
authors.lastname: Malekzadeh
authors.forename: Farzaneh
authors.initials: F
authors.lastname: Sabahi
mesh_terms: D005838:Genotype; D016174:Hepacivirus / Q000235:genetics / Q000302:isolation & purification*; D006526:Hepatitis C / Q000175:diagnosis / Q000821:virology*; D006801:Humans; D012367:RNA, Viral / Q000097:blood*; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D019562:Viral Load / Q000379:methods*
chemical_list: D012367:RNA, Viral