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We also present a method for independently quantifying AF system performance using 2D autocorrelation analysis of astigmatic fluorescent nanobead images. We applied this method to evaluate both this new LED\u2010based\n                    <jats:italic>openAF<\/jats:italic>\n                    module and our previous SLD\u2010based module implemented on a microscope with a 100\u00a0\u00d7\u00a01.4 numerical aperture oil\u2010immersion objective lens. 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