Lentiviral vectors (LVs) are widely used for the generation of gene-modified cell therapies with their main applications in immuno-oncology and rare diseases.  Yet, the variability of the transduction process may affect the consistency of the manufacturing process causing product heterogeneity. Therefore,  controlling the integration of LVs into the host genome is critical to mitigate a risk for insertional mutagenesis.  We have previously developed a single-cell vector copy number (VCN) assay which allowed for an in-depth measurement of the heterogeneity of vector transduction, enhancing on a population VCN analysis. However,  the workflow currently used is limited by a labour-intensive droplet digital PCR (ddPCR) step, relying on several 3-plex ddPCR reactions to quantify 5 different targets in transduced single cells. In this study, we demonstrate how the ddPCR analytical burden  can be streamlined with a high-order multiplexing strategy, whereby 5 targets are measured simultaneously in one single ddPCR reaction using ratio-based mixing of channel1 and channel2 fluorochrome-conjugated probes. This method improvement allows for faster  and cheaper single cell VCN analysis, unlocking the potential application of this assay to a larger number of samples or higher number of single cells, enabling greater monitoring of the integration of LVs in cell therapy products.   Simplification of analytical assays for the characterisation of complex cell therapy products will facilitate the adoption of novel, enhanced assays in the field, which in turn will help the design and manufacturing of safer, higher quality products.