Combinatorial effects of three ions, namely silicate (Si), calcium (Ca), and magnesium (Mg) ions, on the adhesion and proliferation of MC3T3‐E1 mouse osteoblast‐like cells were evaluated. The cells were cultured in single‐, dual‐, or triple‐ion‐conditioned culture media with systematically changed ion concentrations. The ranges of Si, Ca, and Mg ion concentrations were set as 10–70, 80–400, and 25–500 ppm, respectively. The numbers of adherent live cells were measured after culturing for 3 h and for 1, 3, and 5 days to examine cell adhesion and proliferation, respectively. Mg ions predominantly enhanced cell adhesion in both the dual‐ion (xSi‐zMg and yCa‐zMg) and triple‐ion (xSi‐yCa‐zMg) systems but had no effect when they acted individually in the single‐ion system. Conversely, Si ions predominantly enhanced cell proliferation in most single‐ and triple‐ion‐conditioned media. Evaluation of the combinatorial effects of the three ions on cell adhesion and proliferation revealed that the dual‐ and triple‐ion‐conditioned media mainly conferred synergistic effects on adhesion but antagonistic effects on proliferation. © 2019 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 107A: 1042–1051, 2019.