Spot estimation accuracy of Shack-Hartmann images and its impact on Airborne Aero-Optic Laboratory (AAOL) wavefront statistics are addressed. A study is conducted of an individual spot simulated using a double sinc function under varying degrees of additive non-zero mean Gaussian noise within a 15×15 pixel area-of-interest. The focus of this paper is two-fold. First, the accuracy of four existing centroiding methods including first moment, convolution, Gaussian, and weighted first moment are compared. It is found that the weighted first moment centroid most accurately estimates spot centers but requires significantly more computational time with respect to the first moment method. Second, three image-processing techniques, including gamma correction, thresholding, and windowing, are analyzed to determine their influence on each centroiding method’s spot estimation accuracy. A fourth order gamma correction significantly reduces spot estimation error for three centroiding methods. The key result is that the accuracy of the first moment centroid with an applied gamma correction is comparable to the weighted first moment without the computational burden. Finally, the first moment centroid with gamma correction and weighted first moment centroid are applied to AAOL flight data. Wavefront statistics are computed and compared to the commonly used first moment centroid.