An artificial insemination instrument of bumblebee queens was firstly developed. This instrument consists of location tube, transfer tube, holding tube, head product, and probe apparatus for reproductive tract etc. This instrument was designed to minimize stress and damage of reproductive tract of bumblebee queens. The regulator handle apparatus in artificial insemination instrument was used the principle of lever, that manipulates easy, accurate and rapid insemination of bumblebee queens. By using this instrument, the insemination rate was over 90%. This instrument will be useful for the breeding and conservation of excellent character of bumblebees.