Shahzamani, Kiana, Sabahi, Farzaneh, Merat, Shahin, Rezvan, Houri, Mirab Samiee, Siamak, Karimi Arzanani, Mohsen, Naghizadeh, Ramin (2008). The use of a sensitive RT-Nested PCR method for detection of Hepatitis C virus. , 10(0), 35-41.
Kiana Shahzamani; Farzaneh Sabahi; Shahin Merat; Houri Rezvan; Siamak Mirab Samiee; Mohsen Karimi Arzanani; Ramin Naghizadeh. "The use of a sensitive RT-Nested PCR method for detection of Hepatitis C virus". , 10, 0, 2008, 35-41.
Shahzamani, Kiana, Sabahi, Farzaneh, Merat, Shahin, Rezvan, Houri, Mirab Samiee, Siamak, Karimi Arzanani, Mohsen, Naghizadeh, Ramin (2008). 'The use of a sensitive RT-Nested PCR method for detection of Hepatitis C virus', , 10(0), pp. 35-41.
Shahzamani, Kiana, Sabahi, Farzaneh, Merat, Shahin, Rezvan, Houri, Mirab Samiee, Siamak, Karimi Arzanani, Mohsen, Naghizadeh, Ramin The use of a sensitive RT-Nested PCR method for detection of Hepatitis C virus. , 2008; 10(0): 35-41.

The use of a sensitive RT-Nested PCR method for detection of Hepatitis C virus

Pathobiology Reserach
Article 4, Volume 10, Issue 0, 2008, Pages 35-41    PDF (460.38 K)
Authors
Kiana Shahzamani1; Farzaneh Sabahi* 1; Shahin Merat2; Houri Rezvan3; Siamak Mirab Samiee4; Mohsen Karimi Arzanani5; Ramin Naghizadeh6
1Department of Virology, School of Medical Sciences, Tarbiat Modares University, Tehran, Iran
2Digestive Disease Center, Shariati Hospital, Medical Sciences University of Tehran, Tehran, Iran
3Iranian Blood Transfusion Organization/ Research Center, Tehran, Iran
4Food and Control Research Laboratories, Tehran, Iran
5Department of Genetics, Karolinska University, Sweden
6Day General Hospital Laboratory, Tehran, Iran
Abstract
Objective: Hepatitis C virus is the major cause of viral hepatitis and its diagnosis in suspected specimens is of great importance. The risk of transfusion- transmitted virus infection is primarily the result of failure in serological screening tests to detect recently infected donors in the pre-seroconversion window period of infection. Therefore, sensitive and accurate diagnosis of HCV prior to antibody production to reduce window period is necessary.
Materials and Methods: In the present study, a sensitive and specific RT-Nested PCR method for detection of a conserved HCV 5'UTR sequence was developed. Two pairs of primers for amplification of the target sequence in two rounds of PCR were selected. The developed RT-Nested PCR assay was performed on HCV-antibody confirmed positive samples as well as negative controls and standard samples. In order to compare the results, One Step RT-PCR kit was used in this study.
Results: 25 HCV-positive plasma samples whose positivity were confirmed by ELISA and Western Blot tests, also as well as 10 fold dilutions of a high viral load plasma sample obtained from a HCV-positive patient as standard samples and 25 negative control plasmas from healthy blood donors were collected and tested by this assay. In all of positive samples a 175bp band was observed on agarose gel electrophoresis, but no band could be detected in negative control plasma. Results from developed RT-PCR assay and One Step RT-PCR kit showed a good correlation.
Conclusion: According to the results of this study, the developed RT-Nested PCR assay has a good sensitivity and specificity for diagnosis of HCV infection. It has the advantage of viral genome detection prior to seroconversion and can be used to detect HCV infection during window period
Keywords
RT-PCR; HCV; 5'UTR region؛ Nested PCR؛ RT-PCR
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