Design of functional cationic microgels as conjugation scaffolds

Abstract

We present the development and detailed characterization of a range of amine functionalized microgels for utilization in conjugation reactions. Cationic N-isopropylmethacrylamide (NIPMAm) based microgels were synthesized through copolymerization with a primary amine containing monomer, N-(3-aminopropyl)methacrylamide hydrochloride (APMA). A range of synthesis conditions and monomer feed ratios generated microgels of diverse architectures, in different size ranges and with varying amounts of incorporated primary amines. The efficiency of amine incorporation was quantified using a fluorescence-based assay in order to determine the potential applicability of these particles for controlled bioconjugation reactions. The pH responsivity of all microgels was studied via dynamic light scattering and their height profile was investigated through atomic force microscopy following deposition on a functionalized flat substrate. Tunable resistive pulse sensing was employed to characterize microgels with respect to their number densities and molecular weights. These microgels were then conjugated to two dyes, malachite green and rose bengal with the purpose of investigating accessibility of primary amine groups for conjugation reactions. The microgel–dye constructs were then analyzed for dye content/microgel. Finally, the viability of NIH 3T3 fibroblasts incubated in the presence of varying concentrations of non-conjugated and dye conjugated microgels was studied. Confocal imaging revealed low cellular toxicity under conditions of incubation with low concentrations of microgel–dye conjugates, which is promising for eventual utilization of these constructs in bioimaging applications.