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Register a new userCharacterizing and tracking single colloidal particles with video holographic microscopy
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We use digital holographic microscopy and Mie scattering theory to simultaneously characterize and track individual colloidal particles. Each holographic snapshot provides enough information to measure a colloidal spheres radius and refractive index to within 1%, and simultaneously to measure its three-dimensional position with nanometer in-plane precision and 10 nanometer axial resolution.
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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
We present a new flexible high speed laser scanning confocal microscope and its extension by an astigmatism particle tracking device (APTV). Many standard confocal microscopes use either a single laser beam to scan the sample at relatively low overall frame rate, or many laser beam to simultaneously scan the sample and achieve a high overall frame rate. Single-laser-beam confocal microscope often use a point detector to acquire the image. To achieve high overall frame rates, we use, next to the standard 2D probe scanning unit, a second 2D scan unit projecting the image directly on a 2D CCD-sensor (re-scan configuration). Using only a single laser beam eliminates cross-talk and leads to an imaging quality that is independent of the frame rate with a lateral resolution of 0.235unit{mu m}. The design described here is suitable for high frame rate, i.e., for frame rates well above video rate (full frame) up to a line rate of 32kHz. The dwell time of the laser focus on any spot in the sample (122ns) is significantly shorter than in standard confocal microscopes (in the order of milli or microseconds). This short dwell time reduces phototoxicity and bleaching of fluorescent molecules. The new design opens further flexibility and facilitates coupling to other optical methods. The setup can easily be extended by an APTV device to measure three dimensional dynamics while being able to show high resolution confocal structures. Thus one can use the high resolution confocal information synchronized with an APTV dataset.
Supported lipid bilayers have been studied intensively over the past two decades. In this work, we study the diffusion of single gold nanoparticles (GNPs) with diameter of 20 nm attached to GM1 ganglioside or DOPE lipids at different concentrations in supported DOPC bilayers. The indefinite photostability of GNPs combined with the high sensitivity of interferometric scattering microscopy (iSCAT) allows us to achieve 1.9 nm spatial precision at 1 ms temporal resolution, while maintaining long recording times. Our trajectories visualize strong transient confinements within domains as small as 20 nm, and the statistical analysis of the data reveals multiple mobilities and deviations from normal diffusion. We present a detailed analysis of our findings and provide interpretations regarding the effect of the supporting substrate and GM1 clustering. We also comment on the use of high-speed iSCAT for investigating diffusion of lipids, proteins or viruses in lipid membranes with unprecedented spatial and temporal resolution.
We demonstrate differential dynamic microscopy and particle tracking for the characterization of the spatiotemporal behavior of active Janus colloids in terms of the intermediate scattering function (ISF). We provide an analytical solution for the ISF of the paradigmatic active Brownian particle model and find striking agreement with experimental results from the smallest length scales, where translational diffusion and self-propulsion dominate, up to the largest ones, which probe effective diffusion due to rotational Brownian motion. At intermediate length scales, characteristic oscillations resolve the crossover between directed motion to orientational relaxation and allow us to discriminate active Brownian motion from other reorientation processes, e.g., run-and-tumble motion. A direct comparison to theoretical predictions reliably yields the rotational and translational diffusion coefficients of the particles, the mean and width of their speed distribution, and the temporal evolution of these parameters.
Polarized light microscopy provides high contrast to birefringent specimen and is widely used as a diagnostic tool in pathology. However, polarization microscopy systems typically operate by analyzing images collected from two or more light paths in different states of polarization, which lead to relatively complex optical designs, high system costs or experienced technicians being required. Here, we present a deep learning-based holographic polarization microscope that is capable of obtaining quantitative birefringence retardance and orientation information of specimen from a phase recovered hologram, while only requiring the addition of one polarizer/analyzer pair to an existing holographic imaging system. Using a deep neural network, the reconstructed holographic images from a single state of polarization can be transformed into images equivalent to those captured using a single-shot computational polarized light microscope (SCPLM). Our analysis shows that a trained deep neural network can extract the birefringence information using both the sample specific morphological features as well as the holographic amplitude and phase distribution. To demonstrate the efficacy of this method, we tested it by imaging various birefringent samples including e.g., monosodium urate (MSU) and triamcinolone acetonide (TCA) crystals. Our method achieves similar results to SCPLM both qualitatively and quantitatively, and due to its simpler optical design and significantly larger field-of-view, this method has the potential to expand the access to polarization microscopy and its use for medical diagnosis in resource limited settings.
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