Despite decades of research no vaccine against HIV-1 infection has proven sufficiently protective in humans to pass the mandatory market approval requirements. Mosaic HIV-1 antigens were recently utilized in the vaccine clinical trial, MOSAICO (NCT03964415). This trial involved the administration of adenovirus vectors encoding Gag-Pol and Env mosaic proteins together with soluble Env proteins used in the boost. The first part of this thesis aimed at the investigation, whether the mosaic Gag components of the entailed vaccine are capable to form HIV-1 virus-like particles (VLPs) as observed for naturally occurring Gag sequences upon expression in mammalian cells. The findings demonstrated that these mosaic antigens mediate HIV-1 VLP assembly. The mosaic Env protein was also shown to be incorporated on the surface of these VLPs. A remarkable feature of these Env proteins was the exposure of epitopes for the binding of broadly neutralizing antibodies (bNAbs). The induction of such bNAbs in vaccinees is a desired goal in HIV-1 vaccine development. As VLPs are commonly considered highly immunogenic, these results warrant further testing of the produced mosaic HIV-1 VLPs in preclinical studies as boost components in heterologous vaccine regimens.The second part of this thesis focused on the establishment of a stable HIV-1 VLP producer cell line, which could serve as a tool in optimizing the VLP purification process using a fluorescence-based approach. Gag-GFP HIV-1 VLPs previously facilitated the quantification of VLPs in parallel to the discrimination from cell contaminants such as extracellular vesicles (EVs). This concept was expanded by the introduction of a second reporter protein dTomato harboring a membrane anchor. Resulting VLPs displayed double fluorescent signals while co-released EVs exhibited only red fluorescence. This should allow for the rapid detection, discrimination and quantification of both particle types employing fluorescence detectors. The obtained results provided initial prove of this principle, and thus offer future perspectives for the further development of purification and concentration protocols and methodologies to achieve HIV-1 VLP downstream optimization.