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13 March 2024 Raman imaging for high sensitivity detection of intracellular molecules in cryofixed biological specimens
Kenta Mizushima, Yasuaki Kumamoto, Shoko Tamura, Masahito Yamanaka, Kentaro Mochizuki, Menglu Li, Syusuke Egoshi, Kosuke Dodo, Yasunori Harada, Nicholas I. Smith, Mikiko Sodeoka, Hideo Tanaka, Katsumasa Fujita
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Proceedings Volume PC12855, Advanced Chemical Microscopy for Life Science and Translational Medicine 2024; PC1285519 (2024) https://doi.org/10.1117/12.3000359
Event: SPIE BiOS, 2024, San Francisco, California, United States
Abstract
Raman microscopy enables us to obtain molecular information in biological samples but has suffered from low signal-to-noise ratio (SNR) due to low of Raman scattering cross-section. Here we developed a cryo-stat equipped Raman microscope for low temperature measurement, allowing long time accumulation of Raman signals. We confirmed the SNR improvement in Raman imaging of cryofixed HeLa cells without photodamage under long time observation at low temperature. The reduction of photobleaching in resonant Raman scattering of carotenoid and cytochrome significantly increases the SNR, demonstrated by 7-color high SNR Raman imaging with multiple Raman tags, including EdU, MitoBADY, and alkyne-tagged Coenzyme Q (AltQ2). AltQ2 is a mobile small molecule that cannot be fixed by chemical fixation.
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Kenta Mizushima, Yasuaki Kumamoto, Shoko Tamura, Masahito Yamanaka, Kentaro Mochizuki, Menglu Li, Syusuke Egoshi, Kosuke Dodo, Yasunori Harada, Nicholas I. Smith, Mikiko Sodeoka, Hideo Tanaka, and Katsumasa Fujita "Raman imaging for high sensitivity detection of intracellular molecules in cryofixed biological specimens", Proc. SPIE PC12855, Advanced Chemical Microscopy for Life Science and Translational Medicine 2024, PC1285519 (13 March 2024); https://doi.org/10.1117/12.3000359
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KEYWORDS
Raman spectroscopy
Biological imaging
Biological samples
Molecules
Signal to noise ratio
Molecular biology
Cryogenics
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