['To evaluate the intersession repeatability of retinal thickness measurements in patients with diabetic macular edema (DME) using the Heidelberg Spectralis optical coherence tomography (OCT) algorithm and a publicly available, three-dimensional graph search-based multilayer OCT segmentation algorithm, the Iowa Reference Algorithm.', 'Thirty eyes from 21 patients diagnosed with clinically significant DME were included and underwent consecutive, registered macula-centered spectral-domain optical coherence scans (Heidelberg Spectralis). The OCT scans were segmented into separate surfaces, and the average thickness between internal limiting membrane and outer retinal pigment epithelium complex surfaces was determined using the Iowa Reference Algorithm. Variability between paired scans was analyzed and compared with the retinal thickness obtained from the manufacturer-supplied Spectralis software.', 'The coefficient of repeatability (variation) for central macular thickness using the Iowa Reference Algorithm was 5.26 μm (0.62% [95% confidence interval (CI), 0.43-0.71]), while for the Spectralis algorithm this was 6.84 μm (0.81% [95% CI, 0.55-0.92]). When the central 3 mm was analyzed, the coefficient of repeatability (variation) was 2.46 μm (0.31% [95% CI, 0.23-0.38]) for the Iowa Reference Algorithm and 4.23 μm (0.53% [95% CI, 0.39-0.65]) for the Spectralis software.'] Is reproducibility of diabetic macular edema estimates from SD-OCT affected by the choice of image analysis algorithm?