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    In-vivo Raman spectroscopy: from basics to applications
    (Bellingham, Wash. : SPIE, 2018) Cordero, Eliana; Latka, Ines; Matthäus, Christian; Schie, Iwan W.; Popp, Jürgen
    For more than two decades, Raman spectroscopy has found widespread use in biological and medical applications. The instrumentation and the statistical evaluation procedures have matured, enabling the lengthy transition from ex-vivo demonstration to in-vivo examinations. This transition goes hand-in-hand with many technological developments and tightly bound requirements for a successful implementation in a clinical environment, which are often difficult to assess for novice scientists in the field. This review outlines the required instrumentation and instrumentation parameters, designs, and developments of fiber optic probes for the in-vivo applications in a clinical setting. It aims at providing an overview of contemporary technology and clinical trials and attempts to identify future developments necessary to bring the emerging technology to the clinical end users. A comprehensive overview of in-vivo applications of fiber optic Raman probes to characterize different tissue and disease types is also given.
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    FLIM data analysis based on Laguerre polynomial decomposition and machine-learning
    (Bellingham, Wash. : SPIE, 2021) Guo, Shuxia; Silge, Anja; Bae, Hyeonsoo; Tolstik, Tatiana; Meyer, Tobias; Matziolis, Georg; Schmitt, Michael; Popp, Jürgen; Bocklitz, Thomas
    Significance: The potential of fluorescence lifetime imaging microscopy (FLIM) is recently being recognized, especially in biological studies. However, FLIM does not directly measure the lifetimes, rather it records the fluorescence decay traces. The lifetimes and/or abundances have to be estimated from these traces during the phase of data processing. To precisely estimate these parameters is challenging and requires a well-designed computer program. Conventionally employed methods, which are based on curve fitting, are computationally expensive and limited in performance especially for highly noisy FLIM data. The graphical analysis, while free of fit, requires calibration samples for a quantitative analysis. Aim: We propose to extract the lifetimes and abundances directly from the decay traces through machine learning (ML). Approach: The ML-based approach was verified with simulated testing data in which the lifetimes and abundances were known exactly. Thereafter, we compared its performance with the commercial software SPCImage based on datasets measured from biological samples on a time-correlated single photon counting system. We reconstructed the decay traces using the lifetime and abundance values estimated by ML and SPCImage methods and utilized the root-mean-squared-error (RMSE) as marker. Results: The RMSE, which represents the difference between the reconstructed and measured decay traces, was observed to be lower for ML than for SPCImage. In addition, we could demonstrate with a three-component analysis the high potential and flexibility of the ML method to deal with more than two lifetime components.
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    Looking for a perfect match: multimodal combinations of Raman spectroscopy for biomedical applications
    (Bellingham, Wash. : SPIE, 2021) Schie, Iwan; Stiebing, Clara; Popp, Jürgen
    Raman spectroscopy has shown very promising results in medical diagnostics by providing label-free and highly specific molecular information of pathological tissue ex vivo and in vivo. Nevertheless, the high specificity of Raman spectroscopy comes at a price, i.e., low acquisition rate, no direct access to depth information, and limited sampling areas. However, a similar case regarding advantages and disadvantages can also be made for other highly regarded optical modalities, such as optical coherence tomography, autofluorescence imaging and fluorescence spectroscopy, fluorescence lifetime microscopy, second-harmonic generation, and others. While in these modalities the acquisition speed is significantly higher, they have no or only limited molecular specificity and are only sensitive to a small group of molecules. It can be safely stated that a single modality provides only a limited view on a specific aspect of a biological specimen and cannot assess the entire complexity of a sample. To solve this issue, multimodal optical systems, which combine different optical modalities tailored to a particular need, become more and more common in translational research and will be indispensable diagnostic tools in clinical pathology in the near future. These systems can assess different and partially complementary aspects of a sample and provide a distinct set of independent biomarkers. Here, we want to give an overview on the development of multimodal systems that use RS in combination with other optical modalities to improve the diagnostic performance.