Apparatus and method for shortwave infrared photothermal (SWIP) microscopy
Inventors
Cheng, Ji-Xin • Ni, Hongli • Yuan, Yuhao
Assignees
Publication Number
US-12352944-B2
Publication Date
2025-07-08
Expiration Date
2044-10-04
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Abstract
A short-wave infrared photothermal (SWIP) microscopy system and method for vibrational imaging of a sample generates shortwave infrared excitation light probe light. The excitation light and the probe light are combined to generate a combined beam, which is focused to generate a focused combined beam, which is directed onto the sample to obtain a SWIP signal generated by absorption-induced thermo-optic selective heating of the sample. The SWIP signal is collected through an aperture in a condenser and detected.
Core Innovation
The invention provides a short-wave infrared photothermal (SWIP) microscopy system and method designed for vibrational imaging of samples. This system utilizes a source of shortwave infrared excitation light and a source of probe light, which are combined and focused onto the sample. The system detects a SWIP signal generated by absorption-induced thermo-optic selective heating, which causes localized changes in the refractive index of the sample, forming a photothermal lens that alters probe light propagation. The signal is collected through an aperture in a condenser and detected by a photodiode, enabling subcellular spatial resolution and millimeter-level imaging depth even in highly scattering media.
The problem addressed by this invention stems from limitations in conventional vibrational microscopy techniques used for cellular imaging in intact tissue. Existing methods suffer from limited imaging depth due to strong water absorption in the mid-infrared range and tissue scattering effects in visible or near-infrared Raman microscopy. Techniques that reach millimeter depths, such as spatially offset Raman spectroscopy, have poor spatial resolution incapable of monitoring cellular-level activity. Therefore, there exists a gap for vibrational imaging at deep tissue levels with high spatial resolution.
The SWIP microscopy system and method solve these problems by exploiting the shortwave infrared spectral window, which reduces water absorption and tissue scattering relative to mid-infrared and visible wavelengths. By stimulating overtone transitions of carbon-hydrogen bonds with pulsed excitation light and probing the subsequent photothermal lens with continuous wave probe light, the system achieves high sensitivity and spatial resolution. The system can image single 1-micron polymer beads at depths of 800 micrometers in scattering phantoms and resolve intracellular lipids within intact tumor spheroids and thick biological tissues, thus filling the existing gap for deep tissue vibrational imaging with subcellular resolution.
Claims Coverage
There are two independent claims covering a SWIP microscopy system and a SWIP microscopy method, each comprising key inventive features related to generating, combining, focusing excitation and probe lights, obtaining and detecting a SWIP signal.
Short-wave infrared photothermal microscopy system architecture
A system comprising a source of shortwave infrared excitation light, a source of probe light, an optical combining element to generate a combined beam, an objective to focus and direct the beam onto a sample to induce absorption-induced thermo-optic selective heating generating a SWIP signal, a condenser with an aperture to collect the SWIP signal, and a detection element to detect the signal from the condenser.
Short-wave infrared photothermal microscopy method steps
A method comprising generating shortwave infrared excitation light and probe light, combining these lights to form a combined beam, focusing the combined beam onto the sample to obtain a SWIP signal generated by absorption-induced thermo-optic selective heating, collecting the SWIP signal through a condenser aperture, and detecting the SWIP signal from the condenser.
The claims cover the core inventive concepts of a SWIP microscopy system and method that use combined shortwave infrared excitation and probe lights to generate and detect a SWIP signal originating from thermo-optic effects induced by selective absorption in the sample, enabling deep tissue vibrational imaging with high spatial resolution.
Stated Advantages
Enables millimeter-deep tissue vibrational imaging with micron-level lateral resolution.
Significantly higher signal amplitude compared to optically detected photoacoustic signals, enhancing detection sensitivity.
Allows imaging through highly scattering media while maintaining spatial resolution beyond the capability of near-infrared stimulated Raman scattering.
Non-contact optical detection prevents signal loss during propagation and avoids mechanical contact with sensitive samples.
Capability to selectively detect photothermal signals distinct from photoacoustic signals by adjusting focusing conditions.
Fills existing gaps in vibrational imaging by combining deep penetration, subcellular resolution, and high sensitivity.
Potential for in-vivo imaging and improved imaging speed with higher repetition rate lasers.
Offers chemical selectivity compatible with differentiating several molecular species including lipids, proteins, and DNA.
Documented Applications
Imaging intracellular lipids in intact tumor spheroids in vitro, supporting cancer metabolism studies.
High-resolution vibrational imaging of thick biological tissues such as fresh swine liver slices, mouse ear, brain slices, and human breast biopsies.
Potential in-vivo imaging demonstrated with epi-detected SWIP imaging of mouse ear and brain tissue.
Dynamic vibrational imaging in biomedical research, including cancer pathology, drug discovery, and tissue morphology studies without tissue sectioning or labeling.
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