The acceleration of bone regeneration by low-intensity laser irradiation may hold potential benefits in clinical therapy in orthopedics and dentistry. The purpose of this study is to compare the effects of light-emitting diode (LED) and laser on pre-osteoblast MC3T3 proliferation and differentiation. Cells were irradiated with red, infrared, and LED (3 and 5 J/cm²). Lasers had a power density of 1 W/cm² and irradiation time of 2 and 5 s. LED had a power density of 60 mW/cm² and irradiation time of 50 and 83 s. Control group did not receive irradiation. Cell growth was assessed by a colorimetric test (MTT) (24, 48, 72, and 96 h), and cell differentiation was evaluated by alkaline phosphatase (ALP) quantification after growth in osteogenic medium (72 and 96 h and 7 and 14 days). At 24 h, the cell growth was enhanced 3.6 times by LED (5 J/cm²), 6.8 times by red laser (3 J/cm²), and 10.1 times by red laser (5 J/cm²) in relation to control group (p < 0.05). At the other periods, there was no influence of irradiation on cell growth (p > 0.05). The production of ALP was not influenced by irradiation at any period of time (p > 0.05). Low-intensity laser and LED have similar effects on stimulation of cell growth, but no effect on cell differentiation.