This clinical study assessed photobiomodulation (PBM) induced analgesic effects of diode lasers and an LED light source on the dental pulp. Baseline responses to electric pulp testing (EPT) were recorded in 93 healthy unrestored premolar teeth in 26 adults (age range 22–63 years) attending a private dental practice. The teeth were irradiated on buccal and lingual aspects of the crown, by placing the tips on the middle third of the crown of the teeth, on separate days for each of 4 different light sources (660, 808, or 904 nm diode lasers, or a novel multi-wavelength LED light source (700–1100 nm)) using comparable parameters (100 mW, 30 s, 6 J). EPT scores were measured after a further 1-, 2-, 5- and 20-min. Discomfort caused by PBM therapy was recorded using the Wong-Baker scale. EPT changes were tracked over time using repeated measures analysis of variance. Baseline EPT scores were very consistent between different days (linear regression r2 0.9422–0.9648). All PBM devices caused a significant elevation in EPT at 5 min, with an earlier onset at 2 mins for 904 nm and LED. The LED was the only light source that elevated scores at 20 min. Across 2–20 min, when ranked by effectiveness, the greatest EPT elevations were seen for LED, followed by 904 nm, then 660 nm and finally 808 nm. Discomfort during PBM was most common with 904 nm, followed by 808 nm. No discomfort occurred from the LED. Among the light sources utilized, the LED multi-wavelength system demonstrated the largest increase in EPT readings, suggesting its potential as a non-pharmacological alternative for achieving dental analgesia compared to diode lasers.