Abstract
To overcome the physical barriers caused by light diffraction, super-resolution techniques are often applied in fluorescence microscopy. State-of-the-art approaches require specific and often demanding acquisition conditions to achieve adequate levels of both spatial and temporal resolution. Analyzing the stochastic fluctuations of the fluorescent molecules provides a solution to the aforementioned limitations, as sufficiently high spatio-temporal resolution for live-cell imaging can be achieved using common microscopes and conventional fluorescent dyes. Based on this idea, we present COL0RME, a method for covariance-based super-resolution microscopy with intensity estimation, which achieves good spatio-temporal resolution by solving a sparse optimization problem in the covariance domain and discuss automatic parameter selection strategies. The method is composed of two steps: the former where both the emitters' independence and the sparse distribution of the fluorescent molecules are exploited to provide an accurate localization; the latter where real intensity values are estimated given the computed support. The paper is furnished with several numerical results both on synthetic and real fluorescence microscopy images and several comparisons with state-of-the art approaches are provided. Our results show that COL0RME outperforms competing methods exploiting analogously temporal fluctuations; in particular, it achieves better localization, reduces background artifacts, and avoids fine parameter tuning.
Highlights
In the field of fluorescence microscopy, the main factor characterizing the microscope resolution is the limit imposed by the diffraction of light: structures with size smaller than the diffraction barrier cannot be well distinguished nor localized
T=500 (4x zoom), Background estimation result on estimated support using continuous exact relaxation of the l0 norm (CEL0) and l1 regularization, Ground truth (GT) background image. (b)High-background (HB) dataset: Diffraction limited image y = 1
We further performed preliminary comparisons with the Entropy-based Super-resolution Imaging (ESI), 3B and bSOFI approaches using available codes provided by the authors on the web[1], but we did not successfully obtain satisfactory results, so we omit them in the following
Summary
In the field of fluorescence (or, more generally, light) microscopy, the main factor characterizing the microscope resolution is the limit imposed by the diffraction of light: structures with size smaller than the diffraction barrier (typically around 250nm in the lateral direction) cannot be well distinguished nor localized. A large and powerful family of imaging techniques achieving nanometric resolution are the ones often known as Single Molecule Localization Microscopy (SMLM) techniques, see, e.g.(1,2) for a review. Among them, methods such as Photo-Activated Localization Microscopy (PALM)(3) and STochastic Optical Reconstruction Microscopy (STORM)(4) are designed so as to create a superresolved image (achieving around 20nm of resolution) by activating and precisely localizing only a few molecules in each of thousands of acquired frames at a time. (4x zoom), Background estimation result on estimated support using CEL0 and l1 regularization, Ground truth (GT) background image.
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
