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Register a new userSelf-blinking Dyes unlock High-order and Multi-plane Super-resolution Optical Fluctuation Imaging
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Added by
Kristin Gru{\\ss}mayer
Publication date
2020
fields
Physics
and research's language is
English
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Diffraction unlimited super-resolution imaging critically depends on the switching of fluorophores between at least two states, often induced using intense laser light and special buffers. The high illumination power or UV light required for appropriate blinking kinetics is currently hindering live-cell experiments. Recently, so-called self-blinking dyes that switch spontaneously between an open, fluorescent on-state and a closed colorless off-state were introduced. Here we exploit the synergy between super-resolution optical fluctuation imaging (SOFI) and spontaneously switching fluorophores for 2D functional and for volumetric imaging. SOFI tolerates high labeling densities, on-time ratios, and low signal-to-noise by analyzing higher-order statistics of a few hundred to thousand frames of stochastically blinking fluorophores. We demonstrate 2D imaging of fixed cells with a uniform resolution up to 50-60 nm in 6th order SOFI and characterize changing experimental conditions. We extend multiplane cross-correlation analysis to 4th order using biplane and 8-plane volumetric imaging achieving up to 29 (virtual) planes. The low laser excitation intensities needed for self-blinking SOFI are ideal for live-cell imaging. We show proof-of-principal time-resolved imaging by observing slow membrane movements in cells. Self-blinking SOFI provides a route for easy-to-use 2D and 3D high-resolution functional imaging that is robust against artefacts and suitable for live-cell imaging.
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We report an experimental demonstration of a nonclassical imaging mechanism with super-resolving power beyond the Rayleigh limit. When the classical image is completely blurred out due to the use of a small imaging lens, by taking advantage of the intensity fluctuation correlation of thermal light, the demonstrated camera recovered the image of the resolution testing gauge. This method could be adapted to long distance imaging, such as satellite imaging, which requires large diameter camera lenses to achieve high image resolution.
Beam self-cleaning (BSC) in graded-index (GRIN) multimode fibres (MMFs) has been recently reported by different research groups. Driven by the interplay between Kerr effect and beam self-imaging, BSC counteracts random mode coupling, and forces laser beams to recover a quasi-single mode profile at the output of GRIN fibres. Here we show that the associated self-induced spatiotemporal reshaping allows for improving the performances of nonlinear fluorescence microscopy and endoscopy using multimode optical fibres. We experimentally demonstrate that the beam brightness increase, induced by self-cleaning, enables two and three-photon imaging of biological samples with high spatial resolution. Temporal pulse shortening accompanying spatial beam clean-up enhances the output peak power, hence the efficiency of nonlinear imaging. We also show that spatiotemporal supercontinuum generation is well-suited for large-band nonlinear fluorescence imaging in visible and infrared domains. We substantiated our findings by multiphoton fluorescence imaging in both microscopy and endoscopy configurations.
Super-resolution fluorescence microscopy is an important tool in biomedical research for its ability to discern features smaller than the diffraction limit. However, due to its difficult implementation and high cost, the universal application of super-resolution microscopy is not feasible. In this paper, we propose and demonstrate a new kind of super-resolution fluorescence microscopy that can be easily implemented and requires neither additional hardware nor complex post-processing. The microscopy is based on the principle of stepwise optical saturation (SOS), where $M$ steps of raw fluorescence images are linearly combined to generate an image with a $sqrt{M}$-fold increase in resolution compared with conventional diffraction-limited images. For example, linearly combining (scaling and subtracting) two images obtained at regular powers extends resolution by a factor of $1.4$ beyond the diffraction limit. The resolution improvement in SOS microscopy is theoretically infinite but practically is limited by the signal-to-noise ratio. We perform simulations and experimentally demonstrate super-resolution microscopy with both one-photon (confocal) and multiphoton excitation fluorescence. We show that with the multiphoton modality, the SOS microscopy can provide super-resolution imaging deep in scattering samples.
The resolution of optical imaging devices is ultimately limited by the diffraction of light. To circumvent this limit, modern super-resolution microscopy techniques employ active interaction with the object by exploiting its optical nonlinearities, nonclassical properties of the illumination beam, or near-field probing. Thus, they are not applicable whenever such interaction is not possible, for example, in astronomy or non-invasive biological imaging. Far-field, linear-optical super-resolution techniques based on passive analysis of light coming from the object would cover these gaps. In this paper, we present the first proof-of-principle demonstration of such a technique. It works by accessing information about spatial correlations of the image optical field and, hence, about the object itself via measuring projections onto Hermite-Gaussian transverse spatial modes. With a basis of 21 spatial modes in both transverse dimensions, we perform two-dimensional imaging with twofold resolution enhancement beyond the diffraction limit.
It has been shown that negative refraction makes a perfect lens. However, with little loss, the imaging functionality will be strongly compromised. Later on, it was proved that positive refraction from Maxwells fish-eye lens can also makes a perfect lens. However, strong debating happens on the introduced drain problem at the imaging position. In this work, we for the first time find that a solid immersion Maxwells fish-eye lens could be used for super-resolution imaging. We find that it is due to the perfect focusing and total reflection at the outer interface, such that a super-resolution image is formed at the required position in the air background. This simple mechanism will also be valid for other absolute instruments and more versatile super-imaging systems will be anticipated.
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