Detection of HCV-RNA by real time PCR using SYBRGREEN DYE I
- Clinical Pathology Department, , Faculty of Medicine, Mansoura University, Egypt.
- Biochemistry department, , Faculty of Medicine, Mansoura University, Egypt.
- Internal Medicine department, Faculty of Medicine, Mansoura University, Egypt.
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Hepatitis C is an infectious disease affecting primarily the liver, with about 3–4 million people are infected yearly, and more than 350,000 people die per year from hepatitis C-related diseases .There is a number of diagnostic tests for hepatitis C including: recombinant immunoblot assay, HCV antibody enzyme immunoassay or ELISA and quantitative HCV RNA polymerase chain reaction (PCR). Several approaches have been employed to detect PCR products. The most popular using TaqMan probe and the other using fluorescent dye SYBR GREEN I. Unlike TaqMan florescent probes, SYBR GREEN I technology method does not require a probe for the qualitative detection and the PCR product is monitored continuously by SYBR GREEN I dye during one step PCR. Aim of work: Quantitative detection of HCV RNA by a simple, rapid, low cost and sensitive real time PCR using SYBR GREEN I dye and compare it with TaqMan technology. Subjects & Methods: The study recruited 150 HCV antibody positive patients. They were (92 males &58 females) with their ages ranging from (21:59 years). Quantitative HCV RNA was performed by both the ordinary TaqMan method as well as SYBR GREEN based method. Results: HCV RNA was detected by Taqman probe in 138 (92%) of total cases with a median of 3.98 × 105 while, it was detected by SYBR GREEN probe in 134 (89.3%) with median of 3.1 × 105. In addition to this, there was a substantial (good) agreement between both methods regarding qualitative assessment (Weighted ?=0.642). Furthermore, stratification of expression results into negative, Low, moderate and high quantities, revealed moderate agreement between both methods as regard quantitative assessment (Weighted ?=0.503). In addition, At <12 IU/mL level, sensitivity of SYBR GREEN I method was 94.4%, specificity was 66.7%, PPV was 97%, NPV was 50% and accuracy was 92%. Conclusion: detection of HCV-RNA by syber green method could be used as a good screening test for qualitative detection especially in blood donor screening.
[Mohammad Hosam Eldeen Zaghloul, Ehsan Mohammad Rizk, Amany Ramadan Elsayed, Neven Farouk Abbas (2015); Detection of HCV-RNA by real time PCR using SYBRGREEN DYE I Int. J. of Adv. Res. 3 (Jun). 1450-1458] (ISSN 2320-5407). www.journalijar.com
