Abstract: Despite the progress made in the prevention of transfusion transmitted infections, transmission of Hepatitis B Virus (HBV) from donors with Occult Hepatitis B virus (OHB) still poses a challenge to the safety of blood donation. The presence of OHB in anti-hepatitis B core antigen (anti-HBc) positive/anti-hepatitis B surface antigen (anti-HBs) negative blood donors has recently been documented worldwide however, its prevalence in anti-HBc positive/anti-HBs positive donors remains obscure. Therefore, this study was aimed to evaluate the presence of OHB among anti-HBc/anti-HBs-positive blood donors. To meet these objectives, a total of 1000 serum samples of consenting blood donors negative for Hepatitis B surface Antigen (HBsAg) were collected at the Hera’a General Hospital, Makkah Al-Mukaramah, Saudi Arabia and screened for anti-HBs and anti-HBc antibodies. According to their serological status, the samples were classified into anti-HBc negative/anti-HBs negative; anti-HBc positive/anti-HBs negative and anti-HBc positive/anti-HBs positive donors. Samples that showed positive reactivity for both anti-HBc/anti-HBs antibodies were then selected for quantitative detection of HBV DNA by using RT-PCR. Results showed that of the 1000 HBsAg-negative donors, 12 (1.2%) were anti-HBc positive/anti-HBs negative and 63 (6.3%) were anti-HBc positive/anti-HBs positive. Additionally, among these 63 anti-HBc positive/anti-HBs positive blood donor population HBV DNA was detected in two of them (3.2%). In conclusion, the results showed the presence of HBV DNA in the sera of anti-HBc positive/anti-HBs positive blood donors in Saudi Arabia. Overall, these results can raise an important public health issue that an anti-HBc positive/anti-HBs positive serum is not a sign of total HBV eradication and in turn, highlight the need for a stringent and better screening system to prevent post-transfusion HBV infection.