Rapid low-cost detection of hepatitis C virus RNA in HCV-infected patients by real-time RT-PCR using SYBR Green I

• Published in: Archives of Iranian medicine Vol. 14; no. 6; p. 396
• Main Authors: Shahzamani, Kiana, Sabahi, Farzaneh, Merat, Shahin, Sadeghizadeh, Majid, Lashkarian, Hamed Esmail, Rezvan, Houri, Samiee, Siamak Mirab, Arzanani, Mohsen Karimi, Jabbari, Hossain, Malekzadeh, Reza
• Format: Journal Article
• Language: English
• Published: Iran Academy of Medical Sciences of I.R. Iran 01-11-2011
• Subjects: Fluorescent Dyes; Hepacivirus - genetics; Hepacivirus - isolation & purification; Hepatitis C - blood; Hepatitis C - diagnosis; Humans; Limit of Detection; Organic Chemicals; Real-Time Polymerase Chain Reaction - economics; Real-Time Polymerase Chain Reaction - methods; RNA, Viral - analysis; RNA, Viral - blood; Sensitivity and Specificity; Time Factors
• ISSN: 1029-2977; 1735-3947

We intend to design and validate a low-cost assay for the detection of hepatitis C virus (HCV) RNA using rapid-cycle RT-PCR. The procedure is performed in a closed system with little risk of contamination allowing PCR and product identification to be performed within one or two hours.
A SYBR Green-based real-time RT-PCR for rapid detection of HCV. Amplicon synthesis was monitored continuously by SYBR Green I, which binds to double stranded DNA during PCR. The PCR products were identified by melting curve analysis. Standard sera with known concentrations of HCV RNA and 150 clinical samples were used to validate our assay.
The minimum detection level of our assay was less than 50 IU/mL. The results on 100 plasma samples were comparable with commercial assays.
This method is useful for rapid qualitative detection of HCV infection and particularly suitable for routine diagnostic applications.