Toward quantitative super-resolution microscopy: molecular maps with statistical guarantees.

Abstract

Quantifying the number of molecules from fluorescence microscopy measurements is an important topic in cell biology and medical research. In this work, we present a consecutive algorithm for super-resolution (stimulated emission depletion (STED)) scanning microscopy that provides molecule counts in automatically generated image segments and offers statistical guarantees in form of asymptotic confidence intervals. To this end, we first apply a multiscale scanning procedure on STED microscopy measurements of the sample to obtain a system of significant regions, each of which contains at least one molecule with prescribed uniform probability. This system of regions will typically be highly redundant and consists of rectangular building blocks. To choose an informative but non-redundant subset of more naturally shaped regions, we hybridize our system with the result of a generic segmentation algorithm. The diameter of the segments can be of the order of the resolution of the microscope. Using multiple photon coincidence measurements of the same sample in confocal mode, we are then able to estimate the brightness and number of molecules and give uniform confidence intervals on the molecule counts for each previously constructed segment. In other words, we establish a so-called molecular map with uniform error control. The performance of the algorithm is investigated on simulated and real data.