The effects of imaging system resolution and laser sheet thickness on the measurement of the Batchelor scale were investigated in a single-cylinder optical engine. The Batchelor scale was determined by fitting a model spectrum to the dissipation spectrum that was obtained from fuel tracer planar laser-induced fluorescence (PLIF) images of the in-cylinder scalar field. The imaging system resolution was quantified by measuring the step-response function; the scanning knife edge technique was used to measure the 10-90% clip width of the laser sheet. In these experiments, the spatial resolution varied from a native resolution of 32.0 μm to 137.4 μm, and the laser sheet thickness ranged from 108 μm to 707 μm. Thus, the overall resolution of the imaging system was made to vary by approximately a factor of four in the in-plane dimension and a factor of six in the out-of-plane dimension. The Batchelor scale was found to increase linearly with the laser sheet thickness; a beam half-width of less than six times the effective in-plane resolution was required for 10% accuracy. The in-plane spatial resolution of the imaging system does not need to conform to the Nyquist condition, where at least two pixels are required to measure a given turbulence length scale, when using a spectral analysis method to calculate the Batchelor scale. Rather, the turbulence length scales can be estimated with 10% accuracy at an effective resolution equal to 1.4 times the Batchelor scale. The major developments of this work are two methods to correct the measured Batchelor scale for finite resolution effects. Based on these results and guidelines, researchers can estimate the error of existing measurements in the literature and design experiments to faithfully resolve the turbulence in engine flows.