Objective To study the spatial distribution of free Ca2+ in osteoclast-like cells cultured on glass. Methods To detect the free Ca2+ in osteoclast-like cells, the images were analyzed with image software, using the laser scanning confocal microscope and fluorescent probe. Results At 37 ℃ the free Ca2+ in osteoclast-like cells could be labelled effectively with 10 μmol/L Fluo-3/AM, the intensity of Ca2+ fluorescent signal in the central part was greater than that in the peripheral part and in the same section the signal was not distributed evenly. Conclusion The intensity of Ca2+ fluorescent signal is different among various organellae in osteoclast-like cell, which suggests the osteoclast-like cell modulate its own function through the spatial difference of free Ca2+ concentration.