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Register a new userTransient vibration imaging with time-resolved synthetic holographic confocal microscopy
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We introduce a new modality for dynamic phase imaging in confocal microscopy based on synthetic optical holography. By temporal demultiplexing of the detector signal into a series of holograms, we record time-resolved phase images directly in the time domain at a bandwidth as determined by the photo detector and digitizer. We demonstrate our method by optical imaging of transient vibrations in an atomic force microscope cantilever with 100 ns time resolution, and observe the dynamic deformation of the cantilever surface after excitation with broadband mechanical pulses. Temporal Fourier transform of a single data set acquired in 4.2 minutes yields frequency and mode profile of all excited out-of-plane vibration modes with sub-picometer vertical sensitivity and sub-micrometer lateral resolution. Our method has the potential for transient and spectroscopic vibration imaging of micromechanical systems at nano- and picosecond scale time resolution.
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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.
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.
Terahertz subwavelength imaging aims at developing THz microscopes able to resolve deeply subwavelength features. To improve the spatial resolution beyond the diffraction limit, a current trend is to use various subwavelength probes to convert the near-field to the far-field. These techniques, while offering significant gains in spatial resolution, inherently lack the ability to rapidly obtain THz images due to the necessity of slow pixel-by-pixel raster scan and often suffer from low light throughput. In parallel, in the visible spectral range, several super-resolution imaging techniques were developed that enhance the image resolution by recording and statistically correlating multiple images of an object backlit with light from stochastically blinking fluorophores. Inspired by this methodology, we develop a Super-resolution Orthogonal Deterministic Imaging (SODI) technique and apply it in the THz range. Since there are no natural THz fluorophores, we use optimally designed mask sets brought in proximity with the object as artificial fluorophores. Because we directly control the form of the masks, rather than relying on statistical averages, we aim at employing the smallest possible number of frames. After developing the theoretical basis of SODI, we demonstrate the second-order resolution improvement experimentally using phase masks and binary amplitude masks with only 8 frames. We then numerically show how to extend the SODI technique to higher orders to further improve the resolution. As our formulation is based on the equation of linear imaging and it uses spatial modulation of either the phase or the amplitude of the THz wave, our methodology can be readily adapted for the use with existing phase-sensitive single pixel imaging systems or any amplitude sensitive THz imaging arrays.
We report experimental results on heterodyne holographic microscopy of subwavelength-sized gold particles. The apparatus uses continuous green laser illumination of the metal beads in a total internal reflection configuration for dark-field operation. Detection of the scattered light at the illumination wavelength on a charge-coupled device array detector enables 3D localization of brownian particles in water
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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