Fig. 1.

Half-life extended H11 shows similar binding to mouse CTLA-4 as H11 but longer half-life than H11. a Schematic representation of anti-mouse CTLA-4 VHH (H11) and H11 linked to anti-serum albumin VHH (H11-HLE). b Binding to mCTLA-4 of H11 and H11-HLE and their EC₅₀ values. His-tagged H11 or H11-HLE was incubated with plate-bound mCTLA-4-Fc. Binding to mCTLA-4 was detected by using HRP-conjugated anti-His tag antibodies and tetramethylbenzidine (TMB). Data are represented as OD. Error bars show SD. c Plasma (left) and tumor (right) concentration–time profiles and half-life of H11 (30 mg/kg) or H11-HLE (30 mg/kg) in MC38 tumor-bearing C57BL/6 mice (n = 3) via intravenous administration. C57BL/6 mice were inoculated with 1 × 10⁶ MC38 cells. When the mean tumor volume reached approximately 300–500 mm³, animals were randomized into treatment groups (n = 3/group) and dosing was initiated. At indicated time points after single dosing, mice were euthanized and plasma and tumor tissues were harvested. H11 or H11-HLE was quantified using ELISA using HRP-conjugated anti-His tag antibodies