Objective · To establish a rapid method to evaluate the activity of agonistic antibody using OX40 (tumor necrosis factor receptor superfamily member 4)/FcγR (Fcγ receptor)-humanized mice. Methods · Bone marrow cells from OX40-humanized mice and FcγR-humanized mice were collected and mixed with equal ratio. Then the mixed bone marrow cells were administrated into irradiated wild-type mice through the tail veins. The reconstruction efficiency of the immune system was confirmed by detecting the expression of hOX40 and hFcγR in the immune cells of chimera mice. After the chimera mice were generated successfully, they were used to evaluate the immunostimulatory activity of anti-hOX40 antibodies to CD4+ or CD8+ T cells. The results of flow cytometry were statistically analyzed. The unpaired t-test was used to compare the means between the two groups, and one-way ANOVA was used to compare the means between multiple groups. Results · Flow cytometry analysis showed that wild-type recipient mice were efficiently reconstituted with hFcγR expressing cells and hOX40 expressing cells to generate OX40/FcγR-humanized bone marrow chimera mice. In these mice, B cells and myeloid cells expressed hFcγRs (P<0.05), and T cells expressed hOX40 upon in vitro stimulation (P<0.05). When these mice were used to evaluate the immunostimulatory activity of anti-hOX40 antibody, significant expressions of IFN-γ and hOX40 were observed (P<0.05). Conclusion · OX40/FcγR-humanized bone marrow chimera mice are generated based on hFcγR expressing cells and hOX40 expressing cells, suggesting a rapid method to build a mouse model with both hFcγR and hOX40 expression. These mice are suitable for evaluating the immunostimulatory activity of agonistic human anti-hOX40 antibodies.