Hepatitis B virus (HBV) infection leaves serological markers that indicate prior contact with the virus or an occult infection. The aim of this study was to determine the profile of HBV markers of individuals from a region of low HBV endemicity excluded from blood donation due to the presence of total anti-HBc antibodies but who tested negative for HBV surface antigen (HBsAg). The profile of 244 subjects exclusively anti-total hepatitis B core (anti-total HBc) reactive was determined. Markers such as anti-HBsAg, HBeAg (HBV core antigen) and anti-HBe (antibody to hepatitis Be antigen) were determined, and HB viral DNA (HBV-DNA) was identified by PCR analysis. Overall, 85.7% of the subjects had an anti-HBs (hepatitis B surface antigen antibody) titer higher than 10 IU/L; 154 samples were negative for HBeAg and 66.5% were reactive for anti-HBe. All samples were negative for HBV-DNA (n = 241). We found a significant association between the anti-HBe and anti-HBs titers (P = 0.026). Most individuals from a low endemic area for HBV have shown anti-HBs titers that confer immunity against HBV, even though they were negative for HBV-DNA. In the light of a shortage of donated blood worldwide, we suggest that special measures should be considered that would facilitate the possible use of these rejected blood units in areas of low endemicity for HBV.