Label-free vibrational imaging by stimulated Raman scattering (SRS) provides unprecedented insight into real-time chemical distributions. Specifically, SRS in the fingerprint region (400–1800 cm−1) can resolve multiple chemicals in a complex bio-environment. However, due to the intrinsic weak Raman cross-sections and the lack of ultrafast spectral acquisition schemes with high spectral fidelity, SRS in the fingerprint region is not viable for studying living cells or large-scale tissue samples. Here, we report a fingerprint spectroscopic SRS platform that acquires a distortion-free SRS spectrum at 10 cm−1 spectral resolution within 20 µs using a polygon scanner. Meanwhile, we significantly improve the signal-to-noise ratio by employing a spatial-spectral residual learning network, reaching a level comparable to that with 100 times integration. Collectively, our system enables high-speed vibrational spectroscopic imaging of multiple biomolecules in samples ranging from a single live microbe to a tissue slice.
Here we demonstrate the use of second harmonic generation (SHG) microscopy-guided synchrotron powder X-ray diffraction (PXRD) for the detection of trace crystalline active pharmaceutical ingredients in a common polymer blend. The combined instrument is capable of detecting 100 ppm crystalline ritonavir in an amorphous hydroxypropyl methylcellulose matrix with a high signal-to-noise ratio (>5000). The high spatial resolution afforded by SHG microscopy allows for the use of a minibeam collimator to reduce the total volume of material probed by synchrotron PXRD. The reduction in probed volume results in reduced background from amorphous material. The ability to detect low crystalline loading has the potential to improve measurements in the formulation pipeline for pharmaceutical solid dispersions, for which even trace quantities of crystalline active ingredients can negatively impact the stability and bioavailability of the final drug product.
CH-π interactions have been cited as an important contributor to carbohydrate recognition. To determine whether N-heterocycles form stronger CH-π interactions, the interactions of methyl ether groups with heterocyclic and nonheterocyclic aromatic surfaces were studied. Both experimental and computational experiments found that N-heterocyclic aromatic surfaces formed stronger interactions. This enhancement was attributed to attractive dipole-dipole interactions between the methyl ether C-O bond and the N-heterocyclic aromatic dipole.
A Mueller tensor mathematical framework was applied for predicting and interpreting the second harmonic generation (SHG) produced with an unpolarized fundamental beam. In deep tissue imaging through SHG and multiphoton fluorescence, partial or complete depolarization of the incident light complicates polarization analysis. The proposed framework has the distinct advantage of seamlessly merging the purely polarized theory based on the Jones or Cartesian susceptibility tensors with a more general Mueller tensor framework capable of handling partial depolarized fundamental and/or SHG produced. The predictions of the model are in excellent agreement with experimental measurements of z-cut quartz and mouse tail tendon obtained with polarized and depolarized incident light. The polarization-dependent SHG produced with unpolarized fundamental allowed determination of collagen fiber orientation in agreement with orthogonal methods based on image analysis. This method has the distinct advantage of being immune to birefringence or depolarization of the fundamental beam for structural analysis of tissues.
Sample-scan phase contrast imaging was demonstrated by producing and coherently recombining light from a pair of axially offset focal planes. Placing a homogeneous medium in one of the two focal planes enables quantitative phase imaging using only common-path optics, recovering absolute phase without halo or obliqueillumination artifacts. Axially offset foci separated by 70 μm with a 10x objective were produced through polarization wavefront shaping using a matched pair of custom-designed microretarder arrays, compatible with retrofitting into conventional commercial microscopes. Quantitative phase imaging was achieved by two complementary approaches: i) rotation of a half wave plate, and ii) 50 kHz polarization modulation with lock-in amplification for detection.
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