Fig. 4. In vivo immunogenicity of polymer-uricase conjugates.

After three weekly administrations of uricase samples, mice were euthanized on the 21st day, and their sera were harvested for the detection of IgM (A) and IgG (B) specific to uricase or uricase conjugates via ELISA test. The mouse spleen was also harvested for the extraction of splenocytes, which were cultured in the presence of native uricase, ZPS-uricase, NZPS-uricase, or MPC-uricase conjugates for 72 hours and then stained for flow cytometry analysis (C). Summary of the percentage of T_reg phenotype (Foxp3⁺) cells among CD4⁺ CD25⁺ splenocytes (D). Statistical significance was determined using Student’s t test.***P < 0.001. Data are represented as mean ± SEM. OD₄₅₀, optical density at 450 nm.