Single-molecule Ellipticity Estimates Using Second-moments Approach the Theoretical Limit
Authors:Wesley Baxter, Shane Stahlheber
Mentor:Alex Small, Associate Professor of Physics, Cal Poly Pomona
Superresolution localization microscopy requires accurate and precise localization algorithms, which determine the precise positions of molecules within images for the purpose of reconstructing a higher resolution image. We have developed a plugin for ImageJ, an image processing and analysis software package created by the National Institutes of Health, called M2LE. This plugin can localize molecules quickly and distinguish between single-molecule and multiple-molecule images using a shape test that requires only a single iteration. The shape test, which uses the ellipticity of the image to determine whether the image contains a single molecule or multiple molecules, is calculated from the first- and second-moments of the image. These ellipticity estimates were found to approach the theoretical limit, assuming Poisson distributed shot noise. Whether an image is correctly identified as either a single molecule or multiple molecules affects the precision and accuracy of the localizations thereof. Moreover, the total numbers of single-molecule images identified, and multi-molecule image rejected, affect the overall quality and accuracy of the final reconstructed superresolution image.