CMV-CD63-mCherry-FLAG Lentivirus

The CMV-CD63-mCherry-FLAG lentivirus is a versatile, ready-to-use gene delivery system designed for robust and stable expression of a CD63 fusion reporter protein using the widely used cytomegalovirus (CMV) promoter. This system utilizes a replication-deficient, self-inactivating lentiviral backbone, enabling long-term expression in both dividing and non-dividing mammalian cells, including primary cells and traditionally difficult-to-transfect cell lines. CD63 is a tetraspanin protein enriched in late endosomes and exosome membranes. Its fusion with the bright red fluorescent protein mCherry and a compact FLAG epitope tag creates a bimodal reporter protein: enabling real-time optical visualization combined with antibody-based detection and enrichment. This combination supports high-contrast live-cell imaging and sensitive biochemical analyses such as Western blotting, immunofluorescence, immunoprecipitation, and affinity capture using anti-FLAG reagents.

This lentiviral reporter is ideally suited for applications in exosome and extracellular vesicle (EV) research, endolysosomal trafficking, and membrane biology. In live-cell and time-lapse microscopy, CD63-mCherry allows dynamic visualization of multivesicular bodies, vesicle budding and release, and co-localization with endosomal markers, thus dissecting pathway dynamics. The FLAG tag adds flexibility for orthogonal detection, facilitating immunocapture of CD63-positive vesicles for downstream proteomics, RNA profiling, or functional uptake studies in recipient cells. Its stable expression supports comparative experiments across clones, conditions, or time points, enabling quantitative assessment of factors regulating EV biogenesis, cargo sorting, secretion, and intercellular communication. In co-culture systems, the fluorescent signal provides a visual readout of vesicle exchange and uptake, while FLAG-based enrichment methods allow precise recovery of donor-derived extracellular vesicles (EVs) for mechanistic studies.