Polymer hydrogel capsules comprised of poly(methacrylic acid) chains and cross-linked via disulfide linkages were investigated for their cytotoxicity and mechanism of internalization in a variety of mammalian cells. The capsules were internalized by all the tested cell lines which differed in their morphology and function and over short to medium term (24 h) revealed no reduction in viability and metabolic activity of cells. The mechanism of capsule uptake was analyzed using inhibitors of various cellular entry pathways. Of these, blocking the clathrin-mediated endocytotic pathway resulted in a statistically significant reduction in capsule uptake, suggesting this was the predominant pathway of capsule entry in these cell lines. The uptake of solid particles with similar surface chemistry was not significantly decreased by the inhibitor of the clathrin-mediated pathway, which suggested that softness and concomitant flexibility of the hydrogel capsules were factors governing the entry mechanism. This work represents the first systematic study of the interaction of polymer hydrogel capsules with mammalian cells and provides essential information for the application of these capsules in biomedicine.