Photobiomodulation and laser acupuncture are pain management treatments, offering safe, non-invasive options for patients seeking alternatives to conventional treatments. Therefore, the aim of this research is to evaluate the effects of low-level laser therapy at acupuncture points on pain mechanisms caused by orthodontic separators. For the study, 162 healthy adult volunteers over the age of 18 were selected. Orthodontic separators were installed to stimulate pain. The volunteers were divided into three groups: Group 1 received diode laser (808 nm, 100 mW) at acupuncture points for 40 seconds (4 J energy), Group 2 received diode laser (808 nm, 100 mW) on molars with orthodontic separators for 40 seconds (4 J energy), and Group 3 received a placebo at acupuncture points. The acupuncture points used were: LI4 (Hegu), EX2 (Yintang), LI6 (Pianli), LI11 (Quchi), and LU7 (Lieque). A modified Visual Analogue Scale (VAS) was used to assess pain levels. To evaluate differences in pain intensity between groups and over time, the Friedman and Kruskal-Wallis tests were used. Statistical analysis confirmed a significant difference in pain intensity between groups over time (p < 0.001). No significant difference in pain perception was observed between genders (p = 0.943). The difference between the groups that received laser therapy at acupuncture points and local points was not statistically significant (p = 0.272). It can be concluded that laser therapy at acupuncture points yields results similar to laser therapy for pain induced by the installation of orthodontic separators, with both being more effective than placebo treatment.