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Quality Control Maps: real-time quantitative quality control of single-molecule localization microscopy data
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Edité par CCSD ; Biophysical Society -
International audience. Single molecule localization microscopy (SMLM) has revolutionized the understanding of cellularorganization by reconstructing informative images with quantifiable spatial distributions of moleculesfar beyond the optical diffraction limit. Much effort has been devoted to optimizing localizationaccuracy. One such approach is the assessment of SMLM data quality in real-time, rather than afterlengthy post-acquisition analysis, which nevertheless represents a computational challenge.We overcame this difficulty by implementing an innovative mathematical approach we designed todrastically reduce the computational analysis of particle localization. Our Quality Control Maps (QCM)workflow enables a much higher rate of data processing compared to that limited by the frequencyrequired by current cameras. Accordingly, by using an innovative computational approach for thedetection step and an estimator based on a Gaussian model of the point spread function (PSF), subpixelparticle locations and their accuracy can be determined through a straightforward analyticalcalculation, without the need for iterations. As a true parameter-free algorithm, QCM is robust andadaptable to all types of SMLM data, with high speed enabling the real-time calculation of quantitativequality control indicators. Such features are compatible with smart microscopy, the concept of whichdepends on the adjustment of acquisition parameters in real-time according to analytical results.Finally, the offline QCM mode can be used as a tool to evaluate synthetic or previously acquired data,as well as to teach the basic concepts of SMLM.
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