Translation of therapeutic polymeric nanosystems to patients and industry requires simplified, reproducible and scalable methods for assembly and loading. A single-step in-line process based on nanocoprecipitation of oxazoline-siloxane block copolymers in flow-focusing poly(dimethylsiloxane) microfluidics was designed to manufacture injection-ready nanosystems. Nanosystem characteristics could be controlled by copolymer concentration, block length and chemistry, microchannel geometry, flow rate, aqueous/organic flow rate ratio and payload concentration. The well-tolerated nanosystems exhibited differential cell binding and payload delivery and could confer sensitivity to photodynamic therapy to HeLa cancer cells. Such injection-ready nanosystems carrying drugs, diagnostic or functional materials may facilitate translation to clinical application.