In vitro amplification of the nucleic acids (DNA or RNA) is used in the detection of microbial agents and thus in the diagnosis of infectious diseases, as well as in the diagnoses of oncological and genetic disorders and forensic medicine. The aim of the present study was to compare the isolation methods of the nucleic acids of hepatitis B and C viruses, causative agents of the two significant infections worldwide. Conventional isolation methods were compared with the commercial kits that have been used commonly in recent years, in terms of reliability, cost-effectiveness, contamination risk and duration of the testing time. Five standards for the isolation of the viral nucleic acids of both HBV DNA (Fluorion HBV QNP 2.0) and HCV RNA (Fluorion HCV QNP 2.1) were used. The isolations of the viral nucleic acids of HBV and HCV were done with the conventional methods, phenol-chloroform and guanidine thiocyanate, and the commercial kits Roboscreen and NucleoSpin. The resultant viral nucleic acid load was determined with a spectrophotometer (WPA UV 1101, Biotech Photometer), and their amplification was conducted with Real-Time PCR. The results of the assessments revealed that the highest nucleic acid concentration were obtained with the conventional methods, while they exhibited significant drawbacks such as long duration of the testing time, difficulty in application, and higher contamination risk.