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Register a new userCoherent space-gated microscopy: a step towards deep-tissue phase imaging of biological cells
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High-resolution optical microscopy suffers from a low contrast in scattering media where a multiply scattered wave obscures a ballistic wave used for image formation. To extend the imaging depth, various gating operations - confocal, coherence, and polarization gating - have been devised to filter out the multiply scattered wave. However, these gating methods are imperfect as they all act on the detection plane located outside a scattering medium. Here, we present a new gating scheme, called space gating, that rejects the multiply scattered wave directly at the object plane inside a scattering medium. Specifically, we introduced a 30 $mu$m-wide acoustic focus to the object plane and reconstructed a coherent image only with the ballistic wave modulated by acousto-optic interaction. This method allows us to reject the multiply scattered wave that the existing gating methods cannot filter out and improves the ratio of the ballistic wave to the multiply scattered wave by more than 100 times for a scattering medium more than 20 times thicker than its scattering mean free path. Using the coherent imaging technique based on space gating, we demonstrate the unprecedented imaging capability - phase imaging of optically transparent biological cells fully embedded within a scattering medium - with a spatial resolution of 1.5 $mu$m.
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Label-free imaging approaches seek to simplify and augment histopathologic assessment by replacing the current practice of staining by dyes to visualize tissue morphology with quantitative optical measurements. Quantitative phase imaging (QPI) operates with visible/UV light and thus provides a resolution matched to current practice. Here we introduce and demonstrate confocal QPI for label-free imaging of tissue sections and assess its utility for manual histopathologic inspection. Imaging cancerous and normal adjacent human breast and prostate, we show that tissue structural organization can be resolved with high spatial detail comparable to conventional H&E stains. Our confocal QPI images are found to be free of halo, solving this common problem in QPI. We further describe and apply a virtual imaging system based on Finite-Difference Time-Domain (FDTD) calculations to quantitatively compare confocal with wide-field QPI methods and explore performance limits using numerical tissue phantoms.
We present a technically simple implementation of quantitative phase imaging in confocal microscopy based on synthetic optical holography with sinusoidal-phase reference waves. Using a Mirau interference objective and low-amplitude vertical sample vibration with a piezo-controlled stage, we record synthetic holograms on commercial confocal microscopes (Nikon, model: A1R; Zeiss: model: LSM-880), from which quantitative phase images are reconstructed. We demonstrate our technique by stain-free imaging of cervical (HeLa) and ovarian (ES-2) cancer cells and stem cell (mHAT9a) samples. Our technique has the potential to extend fluorescence imaging applications in confocal microscopy by providing label-free cell finding, monitoring cell morphology, as well as non-perturbing long-time observation of live cells based on quantitative phase contrast.
We propose a new method to image fluorescent objects through turbid media base on Airy beam scanning. This is achieved by using the non-diffractive nature of Airy beams, namely their ability to maintain their shape while penetrating through a highly scattering medium. We show, that our technique can image fluorescent objects immersed in turbid media with higher resolution and signal to noise than confocal imaging. As proof-of-principle, we demonstrate imaging of 1$mu$m sized fluorescent beads through a dense suspension of yeast cells with an attenuation coefficient of 51cm$^{-1}$ at a depth of 90$mu$m. Finally, we demonstrate that our technique can also provide the depth of the imaged object without any additional sectioning.
Nature features a plethora of extraordinary photonic architectures that have been optimized through natural evolution. While numerical optimization is increasingly and successfully used in photonics, it has yet to replicate any of these complex naturally occurring structures. Using evolutionary algorithms directly inspired by natural evolution, we have retrieved emblematic natural photonic structures, indicating how such regular structures might have spontaneously emerged in nature and to which precise optical or fabrication constraints they respond. Comparisons between algorithms show that recombination between individuals inspired by sexual reproduction confers a clear advantage in this context of modular problems and suggest further ways to improve the algorithms. Such an in silico evolution can also suggest original and elegant solutions to practical problems, as illustrated by the design of counter-intuitive anti-reflective coating for solar cells.
Multiphoton microscopy (MPM) has gained enormous popularity over the years for its capacity to provide high resolution images from deep within scattering samples1. However, MPM is generally based on single-point laser-focus scanning, which is intrinsically slow. While imaging speeds as fast as video rate have become routine for 2D planar imaging, such speeds have so far been unattainable for 3D volumetric imaging without severely compromising microscope performance. We demonstrate here 3D volumetric (multiplane) imaging at the same speed as 2D planar (single plane) imaging, with minimal compromise in performance. Specifically, multiple planes are acquired by near-instantaneous axial scanning while maintaining 3D micron-scale resolution. Our technique, called reverberation MPM, is well adapted for large-scale imaging in scattering media with low repetition-rate lasers, and can be implemented with conventional MPM as a simple add-on.
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