Sebastian Dochow

In Vivo Characterization of Atherosclerotic Plaque Depositions by Raman-Probe Spectroscopy and in Vitro Coherent Anti-Stokes Raman Scattering Microscopic Imaging on a Rabbit Model

Visualization as well as characterization of inner arterial plaque depositions is of vital diagnostic interest, especially for the early recognition of vulnerable plaques. Established clinical techniques provide valuable visual information but cannot deliver information about the chemical composition of individual plaques. Here, we employ Raman-probe spectroscopy to characterize the plaque compositions of arterial walls on a rabbit model in vivo, using a miniaturized filtered probe with one excitation and 12 collection fibers integrated in a 1 mm sleeve. Rabbits were treated with a cholesterol-enriched diet. The methodology can improve the efficiency of animal experiments and shows great potential for applications in cardiovascular research. In order to further characterize the plaque depositions visually, coherent anti-Stokes Raman scattering (CARS) microscopy images have been acquired and are compared with the Raman-probe results.

Multicore fiber with integrated fiber Bragg gratings for background-free Raman sensing

In the last years a variety of fiber optic Raman probes emerged, which are only partly suited for in vivo applications. The in vivo capability is often limited by the bulkiness of the probes. The size is associated with the required filtering of the probes, which is necessary due to Raman scattering inside the fibers. We employed in-line fiber Bragg gratings (FBG) as notch filter for the collection path and integrated them in a novel type of Raman probe. Multicore singlemode fibers (MCSMF) were designed and drawn integrating 19 singlemode cores to achieve better collection efficiency. A Raman probe was assembled with one excitation fiber and six MCSMF with inscribed FBGs as collection fibers. The probe was characterized regarding Raman background suppression, collection efficiency, and distance dependence. First Raman measurements on brain tissue are presented.

Combined fiber probe for fluorescence lifetime and Raman spectroscopy

AbstractIn this contribution we present a dual modality fiber optic probe combining fluorescence lifetime imaging (FLIm) and Raman spectroscopy for in vivo endoscopic applications. The presented multi-spectroscopy probe enables efficient excitation and collection of fluorescence lifetime signals for FLIm in the UV/visible wavelength region, as well as of Raman spectra in the near-IR for simultaneous Raman/FLIm imaging. The probe was characterized in terms of its lateral resolution and distance dependency of the Raman and FLIm signals. In addition, the feasibility of the probe for in vivo FLIm and Raman spectral characterization of tissue was demonstrated. Graphical AbstractAn image comparison between FLIm and Raman spectroscopy acquired with the bimodal probe onseveral tissue samples

Classification of Raman spectra of single cells with autofluorescence suppression by wavelength modulated excitation

Wavelength modulated Raman spectroscopy has recently been shown to suppress the fluorescence background generated by the sample and the substrate. Here we apply this technique to collect wavelength modulated Raman spectra from 697 individual cells for a model system of circulating tumour cells that consists of leukocytes from patients blood, acute myeloid leukaemia cells (OCI-AML3), and breast tumour cells BT-20 and MCF-7. We study the classification behaviour of wavelength modulated Raman spectra in comparison to a common background correction method in chemometrics. Classifications using a support vector machine with a radial based kernel function were compared for classical Raman spectra, average Raman spectra of each cell and wavelength modulated Raman spectra. The dataset was divided into 80% training spectra and 20% independent validation spectra. The stability of the classification was tested by performing training and validation 200 times with randomly selected datasets. The results are displayed in box whisker plots. Cell identification based on wavelength modulated Raman spectra gives similar classification rates than classical and averaged Raman spectra with a tendency of reduced accuracies and increased modelling variations. Possible explanations and strategies to further improve the wavelength modulated Raman spectroscopy are discussed.

Diagnosis and screening of cancer tissues by fiber-optic probe Raman spectroscopy

Raman spectroscopy is an emerging biophotonic tool that advanced in recent years due to steady improvements in instrumentation for excitation and collection, and the availability of fiber optic probes. This review describes the principles of fiber optic Raman probes and their applications in cancer research of lung, breast, skin, bladder, brain, cervix, oral cavity and gastrointestinal tract.

Fiber probe for nonlinear imaging applications

Over the past years it had been demonstrated that multimodal imaging combining the nonlinear modalities coherent anti-Stokes Raman scattering (CARS), two-photon excited auto-fluorescence (TPEF) and second harmonic generation (SHG) show a great potential for tissue diagnosis and tumor identification. To extend the applicability of this multimodal imaging approach for in-vivo tissue screening of difficult to access body regions the development of suitable fiber optic probes is required. Here we report about a novel CARS imaging fiber probe consisting of 10,000 coherent light guiding elements preserving the spatial relationship between the entrance and the output of the fiber. Therefore the scanning procedure can be shifted from the distal to the proximal end of the fiber probe and no moving parts or driving current are required to realize in-vivo CARS endoscopy.

Proof of concept of fiber dispersed Raman spectroscopy using superconducting nanowire single-photon detectors.

Due to its high molecular specificity, Raman spectroscopy is a well-established analytical tool. Usually the inelastically scattered Raman light is spectrally dispersed by a spectrometer. Here, we present an alternative method, using an optical fiber as dispersive element. As the group velocity within the fiber is wavelength-dependent, different Raman bands arrive at different times at the detector. In combination with time-correlated single-photon counting, Raman spectra can be measured in the time domain. As detector we implemented a Superconducting Nanowire Single-Photon Detector (SNSPD), which possesses a timing accuracy of about 20 ps. Within this contribution we show first results of Raman spectra measured in the time domain using gradient index fibers of varying length.

Endoscopic fiber probe for nonlinear spectroscopic imaging

We present a compact multimodal fiber probe that enables the simultaneous recording of nonlinear imaging modalities like coherent anti-Stokes Raman scattering (CARS), second harmonic generation (SHG), and two-photon excited auto-fluorescence (TPEF) for biomedical applications. The probe is based on a gradient index lens design and a multi-core fiber supplying the excitation laser light. The multi-core fiber preserves the spatial relationship between the entrance and output; therefore, the laser scanning procedure can be shifted from the distal to the proximal end of the probe. No moving parts or electric power are required in situ. The generated sample signals can be collected in the backward (epi) direction and transferred to a detection setup with a multimode fiber integrated in the probe head. The first CARS/SHG/TPEF multimodal tissue images recorded with the introduced fiber probe will be presented.

Development of a fiber-based Raman probe for clinical diagnostics

A basic problem intrinsic to many clinical diagnostic procedures as well as minimally invasive surgeries is the online invivo classification of tissue. Associated with this problem is the task to determine the boundaries between tissue sections of various degrees of disease progression, which cannot be identified easily. This problem is partly founded in the imaging modalities conventionally used, i.e., white-light endoscopy or fluorescence-based endoscopic imaging. These techniques allow for extracting of only a limited parameter set for judging the physiological or pathological state of tissue. Furthermore, fluorescence-based endoscopy relies on the administration of external labels, which principally disturbs the native tissue. These problems can be circumvented using Raman microspectroscopy as a diagnostic tool. Raman microscopy allows to record vibrational spectra at each sampling point. Therefore the molecular fingerprint of the sample can be deciphered with spatial resolution. It has been shown that Raman spectroscopy in combination with advanced statistical methods can be used to identify and grade tissue samples. However, the conventional approach of judging excised tissue sections by Raman microscopy does not present an approach which can be readily used in the clinics. Here we present our recent progress towards designing a fiber-based Raman probe, which - in perspective - might be incorporated into the working channel of a surgical endoscope. Thereby, it is anticipated to contribute to the clinical routine. We will review the general design principle of such a device and the specific design strategy for our Raman probe in concert with comparative measurements employing a set of home-built and commercially-available devices.

Comparing Raman and fluorescence lifetime spectroscopy from human atherosclerotic lesions using a bimodal probe.

Fluorescence lifetime imaging (FLIm) and Raman spectroscopy are two promising methods to support morphological intravascular imaging techniques with chemical contrast. Both approaches are complementary and may also be used in combination with OCT/IVUS to add chemical specificity to these morphologic intravascular imaging modalities. In this contribution, both modalities were simultaneously acquired from two human coronary specimens using a bimodal probe. A previously trained SVM model was used to interpret the fluorescence lifetime data; integrated band intensities displayed in RGB false color images were used to interpret the Raman data. Both modalities demonstrate unique strengths and weaknesses and these will be discussed in comparison to histologic analyses from the two coronary arteries imaged.