Background Light-emitting diode (LED)-based photobiomodulation is used as an inducer of cell regeneration. Although numerous in vitro and in vivo orthopedic studies have been conducted, the ideal LED wavelength range for tendon healing has not yet been determined. This study, thus, focused on the effects of LED of a 630 nm wavelength on the cell viability, proliferation, and migration of human biceps tendon fibroblast cells. Methods Human tendon fibroblast cell culture was performed using the biceps tendon of patients who had undergone biceps tenodesis. Human biceps tendon fibroblasts from two patients (male, aged 42 and 69 years) were isolated and cultured. The cell type was confirmed by a morphological analysis and using tendon and fibroblast specific markers. They were then split into three groups, with each receiving a different irradiation treatment: no LED treatment (control), 630 nm LED, and 630 nm + 880 nm LED for 20 minutes each. After the LED treatment, cell viability, proliferation, and migration assays were performed, and the results were compared between the groups. Results Twenty-four hours after LED treatment, cell viability and proliferation were significantly increased in the 630 nm LED and 630 nm + 880 nm LED treatment groups compared to that in the control group (p < 0.05). Under the same conditions, compared with the control group, the 630 nm LED alone treatment group showed a 3.06 ± 0.21 times higher cell migration rate (p < 0.05), and the 630 nm + 880 nm LED combination treatment group showed a 2.88 ± 0.20 times higher cell migration rate (p < 0.05) in three-dimensional migration assay. Conclusions In human tendon fibroblast cells, 20 minutes of LED treatment at 630 nm and 630 nm + 880 nm exhibited significant effects on cell proliferation and migration. Our findings suggest the potential of LED therapy as an adjuvant treatment for tendon healing, and hence, further research is warranted to standardize the various parameters to further develop and establish this as a reliable treatment regimen.