The islet transplantation is a promising clinical treatment for type I diabetes. However, a lack of islet donors and enzymatic digestion during islet isolation damaging β-cell survival are problematic. For surviving and functionalizing of β-cells after transplantation, maintenance of cell-to-cell contacts and support of stable construction are required. In previous studies, two processes of making β-cell spheroids for cellular interaction and fabrication of a transplantable carrier were separately conducted. Here, we propose one-step fabrication method of the transplantable construction containing β-cell spheroids by 3D bioprinting process. By hybrid printing with polycaprolactone (PCL), mechanically stable and transplantable constructs were successfully fabricated. In addition, the bioink containing both β-cells and calcium chloride was positioned into the printed alginate channels. This process enabled β-cells to form spheroids within the alginate channels. β-cell spheroids having various diameters of 100 μm to 500 μm could be produced within 1 day by controlling the dispensing rate and cell density. Especially, compared to 2D monolayer culture group and non-spheroid contained group, spheroid contained group showed excellent cell viability, proliferation and glucose stimulated insulin secretion. The proposed method is a new approach to improve islet transplantation, and also could be applied to the other cell spheroid studies.