Single-particle combinatorial multiplexed liposome fusion mediated by DNA

Combinatorial high-throughput methodologies are central for both screening and discovery in synthetic biochemistry and biomedical sciences. They are, however, often reliant on large-scale analyses and thus limited by a long running time and excessive materials cost. We here present a single-particle combinatorial multiplexed liposome fusion mediated by DNA for parallelized multistep and non-deterministic fusion of individual subattolitre nanocontainers. We observed directly the efficient (>93%) and leakage free stochastic fusion sequences for arrays of surface-tethered target liposomes with six freely diffusing populations of cargo liposomes, each functionalized with individual lipidated single-stranded DNA and fluorescently barcoded by a distinct ratio of chromophores. The stochastic fusion resulted in a distinct permutation of fusion sequences for each autonomous nanocontainer. Real-time total internal reflection imaging allowed the direct observation of >16,000 fusions and 566 distinct fusion sequences accurately classified using machine learning. The high-density arrays of surface-tethered target nanocontainers (similar to 42,000 containers per mm(2)) offers entire combinatorial multiplex screens using only picograms of material.

Automated summary

In this study, a single-particle combinatorial multiplexed liposome fusion mediated by DNA is presented, enabling multistep and non-deterministic fusion of nanocontainers. Efficient and leakage-free stochastic fusion sequences were observed for arrays of target liposomes functionalized with individual DNA strands and labeled with distinct chromophores. The method allows high-throughput combinatorial multiplex screens using minimal material.