Prof. Dr. Miguel A. Pleitez (Institute of Biological and Medical Imaging, Bioengineering Center, Helmholtz Munich, Neuherberg, Germany; Chair of Biological Imaging, Central Institute for Translational Cancer Research (TranslaTUM), School of Medicine, Technical University of Munich, Munich, Germany)

Library, A.2.500, Staudtstr. 2

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Abstract:

Mid-infrared (mid-IR) excitation and optoacoustic/optothermal (OA/OT) sensing are excellent examples of highly complementary technologies. Mid-IR absorption excites molecular-specific vibrational transitions that are de-excited in the form of heat, generating OA and OT signals whose intensity primarily depends on efficient heat deposition. The combination of mid-IR excitation and OA/OT detection results in retrieval of high-quality mid-IR spectra (high signal-to-noise ratio) which we apply it for longitudinal assessment of biomolecular composition in living cells and tissues without the need of exogenous labels; i.e., for label-free metabolic imaging. Two different approaches for label-free metabolic imaging are presented: 1) Mid-infraRed Optoacoustic Microscopy (MiROM) and 2) Phase-Shifting Mid-infrared Optothermal Microscopy (PSOM). MiROM uses tightly focused optical excitation with coaxially focused ultrasound detection and chemical-contrast imaging is obtained by raster scanning the sample along the focal plane; simultaneously acquiring OA signals produced at specific molecular vibrations excited by pulsed mid-IR radiation [1]. PSOM is a wide-field chemical-contrast imaging method using pump-probe detection of OT signals by optical phase change due to mid-IR excitation. Chemical-contrast imaging is obtained by the difference in Phase Contrast microscopy images of the sample with and without mid-IR illumination [2]. By MiROM, we can monitor lipid, protein, and carbohydrate dynamics down to the single-cell level with a lateral resolution of ~5 μm in living cells and in thick unprocessed excised tissue. For instance, in living adipocytes, we were able to observe the spatio-temporal distribution of carbohydrates used for triglyceride formation during adipogenesis, to study adipocyte tissue inflammation, and (recently) to access intravital mid-IR signals for non-invasive glucose sensing in mice [1,3,4]. With PSOM, we achieved chemical-contrast in highly confluent live-cell cultures for large field-of-views up to 700 μm in diameter, at high hyperspectral imaging speeds [2]. 
The unique features resulting from the combination of mid-IR excitation with OA/OT sensing have lead to the development of positive-contrast label-free metabolic imaging in living-cells, tissues, and in vivo. This presentation will discuss the basic principles of mid-IR OA and OT sensing for spectroscopy and imaging as well as in the most recent developments on label-free live-cell molecular microscopy, fast analytical histology, and biosensing for longitudinal metabolic research and clinical translation.

References:

[1] Pleitez, M. A.; Khan, A. A.; Soldà, A.; Chmyrov, A.; Reber, J.; Gasparin, F.; Seeger, M. R.; Schätz, B.; Herzig, S.; Scheideler, M., Ntziachristos V., Label-free metabolic imaging by mid-infrared optoacoustic microscopy in living cells, Nat. Biotechnol. 38 (2020) 293-296. https://doi.org/10.1038/s41587-019-0359-9 
[2] Yuan, T.; Riobo,L.; Gasparin, F.; Ntziachristos, V., Pleitez MA., Phase-shifting optothermal microscopy enables live-cell mid-infrared hyperspectral imaging of large cell populations at high confluency, Sci. Adv. 10 (2024) eadj7944. https://doi.org/10.1126/sciadv.adj794
[3] Ko V., Goes M.C., Scheel-Platz L., Yuan T., Chmyrov A., Jüstel D., Ruland J., Ntziachristos V., Keppler S.J., and Pleitez M.A., Fast histological assessment of adipose tissue inflammation by label-free mid-infrared optoacoustic microscopy, npj Imaging 1 (2024). 
[4] Uluç N., Glasl S., Gasparin F., Yuan T., He H., Jüstel D., Pleitez M.A., V. Ntziachristos, Non-invasive measurements of blood glucose levels by time-gating mid-infrared optoacoustic signals, Nat. Metab. (2024). HTTPS://DOI.ORG/10.1038/S42255-024-01016-9.