Characterization of Fluorophore Probe Photostability is critical, as it limits both the retrieval rate and the total quantity of fluorescence information that can be extracted from a specimen. We have developed a reproducible and potentially routine method for quantification of fluorophore photobleaching quantum yields based on a commercially available laser scanning apparatus. In a uniformly distributed sample the scanned beam illuminates a constant number of fluorophores at any time. However at slower scan speeds it dwells longer on certain fluorophores, imparting on them a higher photobleaching probability. The method is applicable for both linear and nonlinear excitation and is inherently consistent with experimental conditions under which laser scanning microscopy is performed. The graph to the right shows a plot of fluorescence attenuation as a function of scan speed for fluorescein in an air-saturated aqueous environment. The data indicate that for every 2000 molecules excited with 780nm MPE, one photobleaches.