Coregistered Spectral Optical Coherence Tomography and Two‐Photon Microscopy for Multimodal Near‐Instantaneous Deep‐Tissue Imaging

Rakhymzhan, Asylkhan; Reuter, Lucie; Raspe, Raphael; Bremer, Daniel; Günther, Robert; Leben, Ruth; Heidelin, Judith; Andresen, Volker; Cheremukhin, Sergey; Schulz‐Hildebrandt, Hinnerk; Bixel, M. Gabriele; Adams, Ralf H.; Radbruch, Helena; Hüttmann, Gereon; Hauser, Anja E.; Niesner, Raluca

doi:10.1002/cyto.a.24012

Cited by 14 publications

(9 citation statements)

References 57 publications

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“…Since most pathophysiological phenomena, independent of the organs they involve, are governed by a complex dynamic interplay of various cellular and non-cellular tissue compartments, simultaneous spectral multiplexing is required in multi-photon imaging of live tissue, in order to elucidate underlying mechanisms. Besides, multi-modal imaging technologies retain the potential to translate in-depth mechanistic understanding of disease pathogenesis into clinical settings, for example, by combining multi-photon microscopy with imaging technologies used in diagnostics, such as magnetic resonance imaging, positron emission tomography, or optical coherence tomography [ 28 ].…”

Section: Discussionmentioning

confidence: 99%

Improvement of the Similarity Spectral Unmixing Approach for Multiplexed Two-Photon Imaging by Linear Dimension Reduction of the Mixing Matrix

Rakhymzhan

Acs

Hauser

et al. 2021

IJMS

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Two-photon microscopy enables monitoring cellular dynamics and communication in complex systems, within a genuine environment, such as living tissues and, even, living organisms. Particularly, its application to understand cellular interactions in the immune system has brought unique insights into pathophysiologic processes in vivo. Simultaneous multiplexed imaging is required to understand the dynamic orchestration of the multiple cellular and non-cellular tissue compartments defining immune responses. Here, we present an improvement of our previously developed method, which allowed us to achieve multiplexed dynamic intravital two-photon imaging, by using a synergistic strategy. This strategy combines a spectrally broad range of fluorophore emissions, a wave-mixing concept for simultaneous excitation of all targeted fluorophores, and an unmixing algorithm based on the calculation of spectral similarities with previously measured fluorophore fingerprints. The improvement of the similarity spectral unmixing algorithm here described is based on dimensionality reduction of the mixing matrix. We demonstrate its superior performance in the correct pixel-based assignment of probes to tissue compartments labeled by single fluorophores with similar spectral fingerprints, as compared to the full-dimensional similarity spectral unmixing approach.

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Section: Discussionmentioning

confidence: 99%

Improvement of the Similarity Spectral Unmixing Approach for Multiplexed Two-Photon Imaging by Linear Dimension Reduction of the Mixing Matrix

Rakhymzhan

Acs

Hauser

et al. 2021

IJMS

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“…The CD19:tdRFP B lymphocyte fate mapping mice 11 , Prx1:tdRFP mesenchymal stromal cells fate mapping mice 77 and Cdh5:tdTomato x Histone:GFP (Cdh5:tdTom) fate mapping mice 61,78 (kind gift of Prof. R. Adams, Dr. M.G. Bixel) used for in vivo imaging were typically 12-24 weeks old females and males.…”

Section: Methodsmentioning

confidence: 99%

In vivooptimized three-photon imaging of intact mouse tibia links plasma cell motility to functional states in the bone marrow

Rakhymzhan,

Fiedler,

Günther

et al. 2023

Preprint

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Intravital multi-photon imaging of the bone marrow is crucial to the study of cellular dynamics, communication with the microenvironment and functions, however, imaging of deep tissue areas is challenging and minimally invasive methods for deep-marrow imaging in intact long bones are needed. We developed a high pulse energy 1650 nm laser prototype, which permits to surpass >100 µm thick cortical bone and to perform three-photon microscopy (3PM) in more than 400 µm depth in the marrow cavity of intact mouse tibiain vivo. Its unique 3 and 4 MHz laser repetition rates allowed us to analyze motility patterns of rare cells over large fields of view deep within the unperturbed marrow. In this way, we found a bi-modal migratory behavior of marrow plasma cells. Besides, the analysis of third harmonics generation (THG) in the tibia identified this signal to be a label-free indicator of the abundance of cellular organelles, in particular the endoplasmic reticulum, reflecting protein biosynthesis capacity. We found that only one third of the plasma cells in the tibia marrow of adult mice have a strong THG signal and, thus, a high protein synthesis capacity, while the other two thirds of plasma cells display a low THG signal. Finally, we identified an inverse link between migratory behavior and THG signal strength in marrow plasma cells. As in these cells, the protein biosynthesis capacity indicated by a strong THG signal is mainly associated with antibody secretion, we could relate motility to functional states of plasma cellsin vivo. Our 3PM method retains the ability to connect cellular dynamics to protein biosynthesis capacity in various marrow cell types beyond plasma cells, as THG is a ubiquitous signal, opening new perspectives on understanding how tissue microenvironment impacts on cellular functions in the bone marrow.

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“…Co-registered OCT and TPEF microscopy has the ability to link specific cellular phenotypes and functions as revealed by TPEF to tissue morphology, thus paving the way between basic research knowledge and clinical observations. 101 SHG provides contrast from non-centrosymmetric molecules such as collagen, which mainly appears in the extracellular matrix as distinct morphological feature. The supramolecular organization can be revealed.…”

Section: Non-linear Optical Microscopy and Spectroscopy: Subcellular Biochemical And Metabolicmentioning

confidence: 99%

Enhanced medical diagnosis for dOCTors: a perspective of optical coherence tomography

et al. 2021

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Significance: After three decades, more than 75,000 publications, tens of companies being involved in its commercialization, and a global market perspective of about USD 1.5 billion in 2023, optical coherence tomography (OCT) has become one of the fastest successfully translated imaging techniques with substantial clinical and economic impacts and acceptance.Aim: Our perspective focuses on disruptive forward-looking innovations and key technologies to further boost OCT performance and therefore enable significantly enhanced medical diagnosis.Approach: A comprehensive review of state-of-the-art accomplishments in OCT has been performed. Results:The most disruptive future OCT innovations include imaging resolution and speed (single-beam raster scanning versus parallelization) improvement, new implementations for dual modality or even multimodality systems, and using endogenous or exogenous contrast in these hybrid OCT systems targeting molecular and metabolic imaging. Aside from OCT angiography, no other functional or contrast enhancing OCT extension has accomplished comparable clinical and commercial impacts. Some more recently developed extensions, e.g., optical coherence elastography, dynamic contrast OCT, optoretinography, and artificial intelligence enhanced OCT are also considered with high potential for the future. In addition, OCT miniaturization for portable, compact, handheld, and/or cost-effective capsule-based OCT applications, home-OCT, and self-OCT systems based on micro-optic assemblies or photonic integrated circuits will revolutionize new applications and availability in the near future. Finally, clinical translation of OCT including medical device regulatory challenges will continue to be absolutely essential.Conclusions: With its exquisite non-invasive, micrometer resolution depth sectioning capability, OCT has especially revolutionized ophthalmic diagnosis and hence is the fastest adopted imaging technology in the history of ophthalmology. Nonetheless, OCT has not been completely exploited and has substantial growth potential-in academics as well as in industry. This applies not only to the ophthalmic application field, but also especially to the original motivation of OCT to enable optical biopsy, i.e., the in situ imaging of tissue microstructure with a resolution approaching that of histology but without the need for tissue excision.

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Coregistered Spectral Optical Coherence Tomography and Two‐Photon Microscopy for Multimodal Near‐Instantaneous Deep‐Tissue Imaging

Cited by 14 publications

References 57 publications

Improvement of the Similarity Spectral Unmixing Approach for Multiplexed Two-Photon Imaging by Linear Dimension Reduction of the Mixing Matrix

Improvement of the Similarity Spectral Unmixing Approach for Multiplexed Two-Photon Imaging by Linear Dimension Reduction of the Mixing Matrix

In vivooptimized three-photon imaging of intact mouse tibia links plasma cell motility to functional states in the bone marrow

Enhanced medical diagnosis for dOCTors: a perspective of optical coherence tomography

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