arXiv subjects

Hongli Ni

Publications and source records attributed to Hongli Ni.

9 recordsLinked to original sources

arXiv2026

Mid-wave infrared photothermal microscopy for molecular and metabolic imaging in deep tissues and spheroids

High-resolution chemical imaging within deep tissues and intact spheroids remains a grand challenge. Here, we introduce mid-wave infrared photothermal (MWIP) microscopy operating in the underexplored 2000-2500 nm spectral window for submicron-resolution molecular and metabolic imaging in intact tumor spheroids and deep tissues. A dark-field photothermal detection scheme significantly suppresses water background and enhances contrast. By accessing strong carbon-hydrogen combination absorptions, a detection limit of 0.12% for dimethyl sulfoxide is achieved, comparable to stimulated Raman scattering microscopy. Depth-resolved imaging of endogenous biomolecules up to 500 micrometers in excised mouse skin and brain tissues is demonstrated. MWIP further enables depth-resolved tracking of transdermal drug transport via carbon-deuterium overtone absorption. Using deuterium metabolic probes, fatty-acid metabolism is imaged at 200 micrometers deep within intact tumor spheroids through carbon-deuterium overtone and combination bands. Collectively, MWIP offers a platform for functional imaging of 3D biological systems in their native environments.

physics.optics↗

arXiv2026

Single-pulse Stimulated Raman Photothermal Microscopy and Direct Visualization of Cholesterol-rich Membrane Domains

By measuring the thermal effects resulting from stimulated Raman excitations, stimulated Raman photothermal (SRP) microscope offers a new access to spatially and temporally resolved Raman signatures across various sample types. Unlike stimulated Raman scattering (SRS) microscopy, SRP is highly compatible with noisy ultrafast laser sources, allowing the use of high peak power, low repetition rate optical parametric amplifier (OPA) lasers to boost sensitivity and imaging speed. Here, we report a single pulse SRP (spSRP) microscope system in which SRP signals are induced by individual pairs of laser pulses generated by an OPA laser. Extensive pulse chirping is used to maximize SRS excitation rate and to minimize photodamage. The single-pixel limit of detection (LOD) of spSRP on dimethyl sulfoxide (DMSO) reaches 890 {\mu}M, which is a ~44-fold improvement over SRS microscope. Versatile applications to fungi, cells, and tissues are demonstrated. Live cell spSRP imaging was carried out at speed of 10 frames per second. Particularly, the spSRP system enables direct visualization of cholesterol-rich domains, co-localized with caveolin immunofluorescence, in the plasma membrane of HeLa cells.

physics.optics↗

arXiv2026

Micromolar chemical imaging by high-energy low-photodamage Coherent Anti-stokes Raman Scattering (HELP-CARS)

Coherent anti-Stokes Raman scattering (CARS) microscopy offers label-free chemical imaging capabilities, but its performance is constrained by small Raman scattering cross-section, strong non-resonant background (NRB), and limited signal-to-noise ratio (SNR). Here, we introduce a high-energy, low-photodamage CARS (HELP-CARS) platform designed to overcome these physical limitations. By employing a 1-MHz non-collinear optical parametric amplifier (NOPA) with extensive pulse chirping, HELP-CARS increases the coherent Raman excitation efficiency by ~300 times and improves the signal-to-nonresonant background ratio by 11 times, while inducing negligible damage during live cell imaging. Furthermore, to remove non-independent noise and physically entangled non-resonant background, we incorporate self-supervised deep-learning denoising and background removal based on the Kramers-Kronig relationship, yielding sensitivity improvement by an additional order of magnitude. Together, these advances provide a micromolar imaging sensitivity (160 uM for Dimethyl sulfoxide-d6) corresponding to 1000 molecules in the focal volume. Such high sensitivity enables high-fidelity chemical imaging in both fingerprint and silent windows. Hyperspectral HELP-CARS imaging of deuterated fatty acids allowed first observation of chemical separation with single lipid droplet. Together, HELP-CARS offers a powerful and generalizable approach for ultrasensitive and quantitative vibrational imaging of biological systems.

physics.optics↗

arXiv2025

3D space-variant modal deconvolution with computed point spread functions

Deconvolution is the most widely used aberration correction technique in microscopy, however most techniques assume that the aberrations are the same for each point in the image, which is rarely true. Methods for tracking spatially varying aberrations require burdensome calibration or computation, or require symmetries in the aberration patterns. Here, we expand on existing modal deconvolution methods to demonstrate 3D fluorescence deconvolution in imaging systems that exhibit no simple symmetry. Our method is based on a space-variant generalization of Richardson-Lucy deconvolution that makes use of ZEMAX\textsuperscript{\textregistered}-derived point spread functions without the requirement of guide stars or calibration measurements. We validate the performance of our method by applying it to snapshot multiplane imaging of both bead samples and biological specimens, and show that modal decomposition is a practical solution for deconvolving spatially varying aberrations that do not display clear symmetries.

physics.optics↗

arXiv2025

Fiber laser based stimulated Raman photothermal microscopy with long working distance optics

Stimulated Raman scattering (SRS) microscopy is a highly sensitive chemical imaging technique. However, the broader application of SRS has been limited by two key challenges: the reliance on low-noise but bulky solid-state laser sources and stringent sample requirements necessitated by high numerical aperture (NA) optics. Here, we present a fiber laser based stimulated Raman photothermal (SRP) microscope that addresses these limitations. While appreciating the portability and compactness of a noisy source, fiber laser SRP enables a two-order-of-magnitude improvement in signal to noise ratio over fiber laser SRS without balance detection. Furthermore, with the use of low NA, long working distance optics for signal collection, SRP expands the allowed sample space from millimeters to centimeters, which diversifies the sample formats to multi-well plates and thick tissues. The sensitivity and imaging depth are further amplified by using urea for both thermal enhancement and tissue clearance. Together, fiber laser SRP microscopy provides a robust, user-friendly platform for diverse applications.

physics.optics↗

arXiv2025

Spatial-offset pump-probe imaging of nonradiative dynamics at optical resolution

Nonradiative photothermal (PT) and photoacoustic (PA) processes have found widespread applications in imaging, stimulation, and therapy. Mapping the generation and propagation of PA and PT waves with resolution is important to elucidate how these fields interact with biological systems. To this end, we introduce spatial offset pump-probe imaging (SOPPI). By spatially offsetting the pump beam and the probe beam, SOPPI can image simultaneously PA and PT wave propagation with nanosecond temporal resolution, micrometer spatial resolution, 65 MHz detection bandwidth, and a sensitivity of 9.9 Pa noise equivalent pressure. We first map the PA and PT evolution from a fiber emitter, and how the wave interacting with a mouse skull and brain slices. SOPPI imaging of PA waves from a tapered fiber with water as an absorber shows a wavelength-dependent generation, evanescent wave generated PA, and back-propagated acoustic Mach Cone. At last, a SOPPI-PACT is developed to reconstruct the pigment distribution inside a zebrafish larva with high precision and signal-to-noise ratio.

physics.optics↗

arXiv2023

Millimeter-deep micron-resolution vibrational imaging by shortwave infrared photothermal microscopy

Deep-tissue chemical imaging plays a vital role in biological and medical applications. Here, we present a shortwave infrared photothermal (SWIP) microscope for millimeter-deep vibrational imaging with sub-micron lateral resolution and nanoparticle detection sensitivity. By pumping the overtone transition of carbon-hydrogen bonds and probing the subsequent photothermal lens with shortwave infrared light, SWIP can obtain chemical contrast from microparticles located millimeter-deep in a highly scattering phantom. By fast digitization on the optically probed signal, the amplitude of photothermal signal is shown to be 63 times larger than that of photoacoustic signal, thus enabling highly sensitive detection of nanoscale objects. SWIP can resolve the intracellular lipids across an intact tumor spheroid and the layered structure in millimeter-thick liver, skin, brain, and breast tissues. Together, SWIP microscopy fills a gap in vibrational imaging with sub-cellular resolution and millimeter-level penetration, which heralds broad potential for life science and clinical applications.

physics.optics↗

arXiv2023

High-content stimulated Raman histology of human breast cancer

Histological examination is crucial for cancer diagnosis, including hematoxylin and eosin (H&E) staining for mapping morphology and immunohistochemistry (IHC) staining for revealing chemical information. Recently developed two-color stimulated Raman histology could bypass the complex tissue processing to mimic H&E-like morphology. Yet, the underlying chemical features are not revealed, compromising the effectiveness of prognostic stratification. Here, we present a high-content stimulated Raman histology (HC-SRH) platform that provides both morphological and chemical information for cancer diagnosis based on un-stained breast tissues. Through spectral unmixing in the C-H vibration window, HC-SRH can map unsaturated lipids, cellular protein, extracellular matrix, saturated lipid, and water in breast tissue. In this way, HC-SRH provides excellent contrast for various tissue components. Considering rapidness is important in clinical trials, we implemented spectral selective sampling to boost the speed of HC-SRH by one order. We also successfully demonstrated the HC-SRH in a clinical-compatible fiber laser-based SRS microscopy. With the widely rapid tuning capability of the advanced fiber laser, a clear chemical contrast of nucleic acid and solid-state ester is shown in the fingerprint result.

q-bio.TO↗

arXiv2021

Multi-window SRS imaging using a rapid widely tunable fiber laser

Spectroscopic stimulated Raman scattering (SRS) imaging has become a useful tool finding a broad range of applications. Yet, wider adoption is hindered by the bulky and environmentally-sensitive solid-state optical parametric oscillator (OPO) in current SRS microscope. Moreover, chemically-informative multi-window SRS imaging across C-H, C-D and fingerprint Raman regions is challenging due to the slow wavelength tuning speed of the solid-state OPO. In this work, we present a multi-window SRS imaging system based on a compact and robust fiber laser with rapid and widely tuning capability. To address the relative intensity noise intrinsic to fiber laser, we implemented auto-balanced detection which enhances the signal-to-noise ratio of stimulated Raman loss imaging by 23 times. We demonstrate high-quality SRS metabolic imaging of fungi, cancer cells, and Caenorhabditis elegans across the C-H, C-D and fingerprint Raman windows. Our re-sults showcase the potential of the compact multi-window SRS system for a broad range of applications.

physics.optics↗