CUMULANT MICROSCOPY

摘要

The invention describes a method and a microscopy system for imaging and analysing stochastically and independently blinking point-like emitters. A multiple-order cumulants analysis in conjunction with an established blinking model enables the extraction of super-resolved environment-related parameter maps, such as molecular state lifetimes, concentration and brightness distributions of the emitter. In addition, such parameter maps can be used to compensate for the non-linear brightness and blinking response of higher-order cumulant images—used for example in Super-resolution Optical Fluctuation Imaging (SOFI)—to generate a balanced image contrast. Structures that otherwise would be masked by brighter regions in the conventional cumulant image become samples using spectral cross-cumulants.

主权项

1. Method for imaging and analysing an object labelled with stochastically and independently blinking point-like emitters, said method comprising the following steps:
Acquiring a time series of images from blinking point-like emitters, Calculating higher-order cumulants from said acquired images, Determining at least the parameter maps for molecular state lifetimes, amplitude and concentration of said emitters by establishing a multi-state blinking model, expressing several cumulant orders as a function of the parameters involved and solving the resulting equations for the parameters, Obtaining a balanced cumulant by spatially deconvolving the cumulant of order n, by taking then the n-th root and by compensating for the temporal weighting factors using the previously determined parameter maps, Displaying said parameter maps and balanced cumulants in a way as to obtain information about the structural and/or functional features of the object reported by said emitters.