Microglia, the resident immune cells of the brain, play essential roles in neuronal development, homeostatic function, and neurodegenerative disease. Human microglia are very different from mouse ones, and thus research of mouse microglia may not always represent the response of human microglia. Further, most research on human microglia is performed in vitro, which does not accurately represent microglia characteristics under in vivo conditions. To address these issues, we developed a simplified method to generate induced pluripotent stem cell-derived human microglia (iPSMG) and transplant them non-invasively into the brain via a transnasal route in immunocompetent mice. We show that by combining pharmacological ON/OFF of a colony stimulating factor 1 receptor antagonist PLX5622 and intranasal transplantation, iPSMG can be non-invasively transplanted into the mouse brain; the transplanted microglia can migrate to each brain site, proliferate and be engrafted for at least 60 d without any immunosuppressants. The relationship between transplanted iPSMG and endogenous mouse MG varies depending on the brain region and transplanted iPSMG mature over a month within the mouse brain. Our method provides a way to non-invasively transplant cells into mouse brain and may therefore be valuable for evaluating how human microglia affect physiological and pathological brain functions.