We report the experimental super-resolution of pairs of point-like fluorescent sources using a modified image inversion interferometer microscope. The technique is inspired by recent developments in the application of quantum parameter estimation theory to semiclassical imaging problems. We find that the image inversion technique requires special polarization filtering to account for the dipolar nature of the emission. Using an azimuthal polarizer, we obtain improvements in the Fisher information of source separation by over an order of magnitude relative to direct imaging. Unlike established super-resolution fluorescence techniques, the method does not require sequential photoswitching/blinking of the fluorophores, and thus could facilitate significant speed-ups for certain biological imaging/tracking tasks. The authors propose and experimentally demonstrate a new route to passive super-resolution fluorescence microscopy inspired by quantum parameter estimation theory, which does not require sequential photoswitching of the fluorophores.
We report the experimental super-resolution of pairs of point-like fluorescent sources using a modified image inversion interferometer microscope. The technique is inspired by recent developments in the application of quantum parameter estimation theory to semiclassical imaging problems. We find that the image inversion technique requires special polarization filtering to account for the dipolar nature of the emission. Using an azimuthal polarizer, we obtain improvements in the Fisher information of source separation by over an order of magnitude relative to direct imaging. Unlike established super-resolution fluorescence techniques, the method does not require sequential photoswitching/blinking of the fluorophores, and thus could facilitate significant speed-ups for certain biological imaging/tracking tasks. The authors propose and experimentally demonstrate a new route to passive super-resolution fluorescence microscopy inspired by quantum parameter estimation theory, which does not require sequential photoswitching of the fluorophores.
Recent theoretical and experimental work has shown that the quantum Fisher information associated with estimating the separation between two optical point sources remains finite at small separations, effectively opening new routes to super-resolution imaging of simultaneously emitting sources. Most studies to date, however, implicitly invoke the scalar approximation, which is not appropriate in the context of high-numerical-aperture microscopy. Utilizing parameter estimation theory, here we consider the estimation of separation between two closely spaced dipole emitters, a commonly employed model for single-molecule optical beacons. We consider two limiting cases: one in which the orientations of the emitters are fixed and equal, and another in which both dipoles freely sample all of orientation space over the course of the measurement. We quantify precision limits using quantum and classical variants of the Fisher information and Cramér-Rao bound. In all cases, the vectorial nature of the emission complicates the analyses, but with appropriate filtering of the collected light in the azimuthal-radial polarization basis, a previously proposed scheme to saturate the quantum Fisher information via image inversion interferometry can be salvaged.
Super-resolution techniques in microscopy are opening new frontiers in biology, and super-resolution for telescope imaging may provide similar benefits for astronomy and space domain awareness. Telescope super-resolution techniques such as multi-plane light conversion and photonic lanterns have been inspired by the ability of mode sorting measurements to extract classical Fisher information closer to the quantum bound than standard telescope imaging. Fisher information analyses for super-resolution imaging systems commonly consider the case of imaging point sources, but often only consider estimating a small subset of the parameters. Furthermore, there has been no demonstration of an estimation algorithm with performance that follows the limit set by the Fisher information when all point source parameters are considered. We provide a framework for defining and analyzing super-resolution using the classical and quantum Fisher information for the multi-parameter estimation problem and demonstrate an algorithm that can achieve the classical Fisher information bound for all the parameters simultaneously.
The classical optical diffraction limit can be overcome by exploiting the quantum properties of light in several theoretical studies; however, they mostly rely on an entangled light source. Recent experiments have demonstrated that quantum properties are preserved in many fluorophores, which makes it possible to add a new dimension of information for super-resolution fluorescence imaging. Here, we developed a statistical quantum coherence model for fluorescence emitters and proposed a new super-resolution method using fluorescence quantum coherence in fluorescence microscopy. In this study, by exploiting a single-photon avalanche detector (SPAD) array with a time-correlated single-photon-counting technique to perform spatial-temporal photon statistics of fluorescence coherence, the subdiffraction-limited spatial separation of emitters is obtained from the determined coherence. We numerically demonstrate an example of two-photon interference from two common fluorophores using an achievable experimental procedure. Our model provides a bridge between the macroscopic partial coherence theory and the microscopic dephasing and spectral diffusion mechanics of emitters. By fully taking advantage of the spatial-temporal fluctuations of the emitted photons as well as coherence, our quantum-enhanced imaging method has the significant potential to improve the resolution of fluorescence microscopy even when the detected signals are weak.
According to Rayleigh's criterion, two incoherent emitters with a separation below the diffraction limit are not resolvable with a conventional fluorescence microscope. One method of superresolution microscopy (SRM) circumvents the diffraction-limited resolution by precisely estimating the position of spatiotemporally independent emitters. However, these methods of SRM techniques are not optimal for estimating the separation of two simultaneously excited emitters. Recently, a number of detection methods based on modal imaging have been developed to achieve the quantum Cramér-Rao lower bound (QCRB) to estimate the separations between two nearby emitters. The QCRB determines the minimum achievable precision for all possible detection methods. Current modal imaging techniques assume a scalar field generated from a point source, such as a distant source from an optical fiber or a pinhole. However, for fluorescently labeled samples, point emitters are single fluorophores that are modeled as dipole emitters and, in practice, are often freely rotating. Dipole radiation must be described by vectorial theory, and the assumption of a scalar field no longer holds. Here, we present a method to numerically calculate the QCRB for measuring the separation of two dipole emitters, incorporating the vectorial theory. Furthermore, we propose a near-quantum optimal detection scheme based on one of the modal imaging techniques, superlocalization by image inversion interferometry (SLIVER), for estimating the separation of two freely rotating dipoles. In the proposed method, we introduce a vortex wave plate before the SLIVER detection to separate the radial and azimuthal components of the dipole radiation. With numerical simulations, we demonstrated that our method achieves nondivergent precision at any separation between two dipole emitters. We investigated several practical effects relevant to experimental measurements in superresolution microscopy, including numerical aperture, detection bandwidth, number of estimation parameters, background, and misalignment on separation estimation. Our proposed measurement provides a near-quantum-limited detection scheme for measuring the separation of two freely rotating dipole emitters, such as fluorescently tagged molecules, which are commonly used in superresolution microscopy.
Multiphoton fluorescence microscopy is now a well-established technique, currently attracting much interest across all fields of biophysics - especially with regard to enhanced focal resolution. The fundamental mechanism behind the technique, identified and understood through the application of quantum theory, reveals new optical polarization features that can be exploited to increase the information content of images from biological samples. In another development, based on a newly discovered, fundamentally related mechanism, it emerges the passage of off-resonant probe laser pulses may characteristically modify the intensity of single-photon fluorescence, and its associated optical polarization behavior. Here, the probe essentially confers optical nonlinearity on the decay transition, affording a means of optical control over the fluorescent emission. Compared to a catalogue of other laser-based techniques widely used in the life sciences, most suffer limitations reflecting the exploitation of specifically lifetime-associated features; the new optical control mechanism promises to be more generally applicable for the determination of kinetic data. Again, there is a prospect of improving spatial resolution, non-intrusively. It is anticipated that tight directionality can be imposed on single-photon fluorescence emission, expediting the development of new imaging applications. In addition, varying the optical frequency of the probe beam can add another dimension to the experimental parameter space. This affords a means of differentiating between molecular species with strongly overlapping fluorescence spectra, on the basis of their differential nonlinear optical properties. Such techniques significantly extend the scope and the precision of spatial and temporal information accessible from fluorescence studies.
The resolution limits of classical spectroscopy can be surpassed by quantum-inspired methods leveraging the information contained in the phase of the complex electromagnetic field. Their counterpart in spatial imaging has been widely discussed and demonstrated; however, the spectral-domain implementations are few and scarce. We experimentally demonstrate a spectroscopic super-resolution method aimed at broadband light (tens to hundreds of GHz), and based on the spectral-domain analog of image inversion interferometry. In a proof-of-principle experiment, we study the paradigmatic problem of estimating a small separation between two incoherent spectral features of equal brightness, with a small number of photons per coherence time. On the grounds of asymptotic estimation theory, more than a two-fold improvement over the spectral direct imaging is demonstrated in terms of required resources (photons) for a given estimator variance. The setup is based on an actively stabilized Mach–Zehnder-type interferometer with electro-optic time lenses and passive spectral dispersers implementing the inversion. As such, the method promises on-chip integration, good scalability, and further applications, e.g., for mode sorting.