A new method for measuring grating groove density is proposed using multiple diffraction orders and one standard wavelength to eliminate the eccentricity effect originated from the rotation of a diffraction grating. Based on the Littman configuration for autocollimation, the groove density can be obtained by measuring the internal angles between the zeroth-order and two nonzeroth-order diffracted beams for one laser wavelength at a certain incident angle. The method was applied to measure the groove density of a plane grating with a nominal groove density of 651 lines/mm, where the mean value of this grating was measured to be 650.76 lines/mm with a relative measurement error / of 3.8×10.