The human cytomegalovirus (hCMV), a double stranded DNA virus of the herpesvirus family, is a ubiquitous human-pathogenic -herpesvirus, capable of inducing a latent infection. In immunocompetent hosts, infections with hCMV often go unnoticed as affected individuals usually suffer from unspecific or no symptoms at all. While hCMV infections can cause hCMV mononucleosis, this syndrome is only of minor clinical importance as it is generally self-limiting and oftentimes does not require antiviral therapy. However, immunocompromised individuals suffer greatly from primary hCMV infections or reactivation, often resulting in life-threatening complications. Congenital hCMV infections are the most common congenital infection, affecting 0,7% of all neonates. Such infections clinically present with cerebral palsy, developmental and cognitive delay, as well as epilepsy, vision loss and deafness. Other groups suffering from hCMV infections are AIDS patients, recipients of solid organ or hematopoietic stem cell transplantations and other conditions which result in immunosuppression. Generally, these individuals suffer from fever, malaise and mild myelosuppression, in addition to tissue-invasive disease focused on the gastrointestinal tract, the liver, the central nervous system and the transplanted allograft. Past efforts to create a vaccine have not been successful, despite being categorized as “most favorable” in 2000 by a report of the Institute of Medicine, estimating annual savings of 820 million USD per year to the US healthcare system through the implementation of a successful hCMV vaccination campaign. Goal of this thesis was the establishment of a cost effective hCMV-specific B-cell Enzyme-linked Immunospot (ELISpot) assay to support vaccine research efforts with a method to easily examine the generation, maintenance and characteristics of hCMV- or vaccine-induced cellular component of the humoral immune system. During the establishment of the ELISpot assay, hCMV-specific spots, suggesting the presence of hCMV-specific B-cells, were observed in samples of individuals previously determined to be hCMV-seronegative by means of a commercial hCMV- Enzyme-linked Immunosorbent Assay (ELISA) kit. Approximately 37,5% of hCMV-seronegative samples displayed reactivity in the hCMV ELISpot assay, which might be the result of false-positive readings of the assay, although the most common causes of such false readings have been ruled out warranting further investigation of the phenomenon. Additionally, we utilized this assay in conjunction with the hCMV-specific ELISA to demonstrated that titers of Immune globulin G (IgG) against hCMV do not correlated with the frequency of circulating hCMV-specific memory B-cells.