Abstract

We pioneer the design of dual-gated microparticles, both responsive to changes in temperature and pH, for stimuli-responsive chromatography targeted at the efficient separation of antibodies. Dual-gated microspheres were synthesized by introducing RAFT-based thiol-terminal block copolymers of poly(N-isopropylacrylamide-b-4-vinylpyridine) (P(NIPAM-b-4VP, 4800 ≤ Mn/Da ≤ 10 000, featuring block length ratios of 29:7, 29:15, and 29:30, respectively) by thiolepoxy driven ligation to the surface of poly(glycidylmethacrylate) (PGMA) microparticles (10−12 μm), whereby the 4-vinylpyridine units within the lateral chain enable protein binding. The switchable protein release abilities of the resulting microparticle resins are demonstrated by adsorption of immunoglobulins at 40 °C and pH 8 and their release at 5 °C or pH 3,respectively. We demonstrate that P(NIPAM29-b-4VP30)-grafted PGMA particles show a maximum adsorption capacity for immunoglobulins of 18.9 mg mL−1 settled resin at 40 °C/pH 8, whereas the adsorption capacity decreased to 7.5 mg mL−1 settled resin at 5 °C while retaining the pH value, allowing the unloading of the chromatographic column by a facile temperature switch. Critically, regeneration of the dual-gated microspheres became possible by lowering the pH to 3.

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