Background: Hepatitis E virus (HEV) is a significant etiological agent of hepatic disease globally, primarily transmitted via the fecal–oral route, often as a consequence of contaminated water or food sources in regions characterized by inadequate sanitation and hygiene infrastructure, such as numerous developing countries. Objectives: This study aims to investigate the seroprevalence of HEV and recognize the risk factors linked to HEV among blood donors in Jordan. Methodology: A cross-sectional study conducted at the blood bank unit of King Hussein Medical Center (KHMC) in Jordan. The study involved collecting 174 blood samples from healthy donors during November 2024. To detect anti-HEV IgG, the serum samples were analyzed using enzyme-linked immunosorbent assays (ELISA). Result: Anti-HEV IgG was detected in 10 out of 174 (5.75%), with a significantly higher rate of HEV negativity (93.2%, P < 0.001). All positive (anti- HEV IgG) samples are males. Amman had the highest percentage of positive samples (55.75%), with no significant association with lifestyle or medical history. Conclusion: The study highlights the potential risk of HEV transmission via blood transfusions. Among Jordanian blood donors, seroprevalence of HEV among blood donors is low, with no significant association with lifestyle or medical history. Further studies with a larger number, and more diverse samples that include both males and females are necessary to investigate transmission routes and risk factors, enhancing public health and transfusion safety.