Background: over the two last decades, much attention has been given to the prevention of transfusion-transmitted viral infections. Given the potential transmission of viruses during the window period, novel non-serology based approaches such as viral nucleic acid testing (NAT) have been established. Objective: a comparative study of different methods to detect transfusion transmitted viral infections. Materials and methods: this cross-sectional study was conducted at the Egyptian National Blood Transfusion Center (NBTC), Giza. The duration of the research was 2 months from June 2018 to august 2018. In our study 1000 donor samples were tested by ELISA and NAT tests for HBV, HCV and HIV. Results: regarding ELISA tests of the three viruses, only 14 (1.4%) samples were finally HBsAg reactive. Finally, reactive HCVAb were 22 (2.2%) samples, while only 4 (0.4%) samples were HIV Ag-Ab finally reactive. We found highly statistical significant difference between initial and final results of ELISA (p<0.001). Regarding NAT testing of the three viruses. HBV: 5 (0.5%) samples were NAT and ELISA reactive, 1 was NAT yield. HCV: 9 (0.9%) samples were NAT and ELISA reactive. HIV one sample (0.1%) was NAT and ELISA reactive, one (0.1%) was NAT yield. Statistical significant difference was found in comparison between NAT and ELISA in HBV, HCV, and HIV p values <0.001, <0.001, and = 0,008 respectively. Conclusion: we concluded that NAT and ELISA are complementary to each other. NAT is an effective method for safeguarding the blood supply.