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Abstract High‐sensitivity chemical imaging offers a window to decipher the molecular orchestra inside a living system. Based on vibrational fingerprint signatures, coherent Raman scattering microscopy provides a label‐free approach to map biomolecules and drug molecules inside a cell. Yet, by near‐infrared (NIR) pulse excitation, the sensitivity is limited to millimolar concentration for endogenous biomolecules. Here, the imaging sensitivity of stimulated Raman scattering (SRS) is significantly boosted for retinoid molecules to 34 micromolar via electronic preresonance in the visible wavelength regime. Retinoids play critical roles in development, immunity, stem cell differentiation, and lipid metabolism. By visible preresonance SRS (VP‐SRS) imaging, retinoid distribution in single embryonic neurons and mouse brain tissues is mapped, retinoid storage in chemoresistant pancreatic and ovarian cancers is revealed, and retinoids stored in protein network and lipid droplets ofCaenorahbditis elegansare identified. These results demonstrate VP‐SRS microscopy as an ultrasensitive label‐free chemical imaging tool and collectively open new opportunities of understanding the function of retinoids in biological systems.
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High-resolution infrared imaging of biological samples with third-order sum-frequency generation microscopy
We studied the use of vibrationally resonant, third-order sum-frequency generation (TSFG) for imaging of biological samples. We found that laser-scanning TSFG provides vibrationally sensitive imaging capabilities of lipid droplets and structures in sectioned tissue samples. Although the contrast is based on the infrared-activity of molecular modes, TSFG images exhibit a high lateral resolution of 0.5µm or better. We observed that the imaging properties of TSFG resemble the imaging properties of coherent anti-Stokes Raman scattering (CARS) microscopy, offering a nonlinear infrared alternative to coherent Raman methods. TSFG microscopy holds promise as a high-resolution imaging technique in the fingerprint region where coherent Raman techniques often provide insufficient sensitivity.
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- Award ID(s):
- 1763272
- PAR ID:
- 10075184
- Publisher / Repository:
- Optical Society of America
- Date Published:
- Journal Name:
- Biomedical Optics Express
- Volume:
- 9
- Issue:
- 10
- ISSN:
- 2156-7085
- Page Range / eLocation ID:
- Article No. 4807
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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