Purpose: Scientific evidence pertaining to the evaluation of trueness of occlusal splints fabricated using different three-dimensional (3D) printers and build orientations compared to subtractive technologies is lacking. Methods: Overall, one hundred and ten occlusal splints were manufactured using two different 3D printers and a dental mill. Five groups of ten were fabricated using the 3D printers at different build orientations (0, 30, 45, 60, and 90 degrees). In addition, a comparison group of ten occlusal splints was subtractively manufactured using a five-axis dental mill. All occlusal splints were scanned and exported as a standard tessellation language file. Analysis was conducted with metrology software with root mean square estimate average positive deviation and average negative deviation used as the measured outcome. Results: The 0 degree printing orientation was the most accurate for printer one with the root mean square value of 0.05 ± 0.01 mm, and 60 degree printing orientation was most accurate for printer two with the RMS value of 0.11 ± 0.01 mm. Subtractively manufactured occlusal splint had significantly higher trueness with the lowest RMS value of 0.03 ± 0.05 mm. Conclusion: Build orientations influence the trueness of additively manufactured occlusal splints while occlusal splints produced by subtractive manufacturing were statistically significantly more accurate.