Methods used in a scientific study to evaluate liver injury markers and analyze serological markers of hepatitis C virus (HCV) infection.
Liver biopsy samples were evaluated using the Ishak scoring system, while serum markers of liver injury and bilirubin were analyzed through routine assays on an automated analyzer. The study also quantified HCV RNA in sera using a specific assay and analyzed the HCV genotypes. To analyze the human IL28B promoter polymorphism at position -3176 (rs12979860), genomic DNA was extracted from peripheral blood using a specific kit and analyzed using the LightMix Kit IL28B.
To analyze gene expression, total RNA was isolated from liver tissue and PBL, and DNase treatment was carried out prior to cDNA synthesis. RealTime HCV RNA and gene expression quantification was performed using specific primer sequences and two reference genes (HPRT and GAPD) selected based on analyses of HCV-infected patients and controls. The relative expression level was determined by measuring the delta cycle threshold value and calculating the fold change.
The specific methods used in a scientific study related to liver injury and HCV infection, including the use of different assays and kits, as well as the concept of gene expression analysis based on primer sequences and reference.
