Real-time molecular imaging of near-surface tissue using Raman spectroscopy.

The steady progress in medical diagnosis and treatment of diseases largely hinges on the steady development and improvement of modern imaging modalities. Raman spectroscopy has attracted increasing attention for clinical applications as it is label-free, non-invasive, and delivers molecular fingerprinting information of a sample. In combination with fiber optic probes, it also allows easy access to different body parts of a patient. However, image acquisition with fiber optic probes is currently not possible. Here, we introduce a fiber optic probe-based Raman imaging system for the real-time molecular virtual reality data visualization of chemical boundaries on a computer screen and the physical world. The approach is developed around a computer vision-based positional tracking system in conjunction with photometric stereo and augmented and mixed chemical reality, enabling molecular imaging and direct visualization of molecular boundaries of three-dimensional surfaces. The proposed approach achieves a spatial resolution of 0.5 mm in the transverse plane and a topology resolution of 0.6 mm, with a spectral sampling frequency of 10 Hz, and can be used to image large tissue areas in a few minutes, making it highly suitable for clinical tissue-boundary demarcation. A variety of applications on biological samples, i.e., distribution of pharmaceutical compounds, brain-tumor phantom, and various types of sarcoma have been characterized, showing that the system enables rapid and intuitive assessment of molecular boundaries.

In-vivo Raman spectroscopy: from basics to applications.

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.

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.

Combined fiber probe for fluorescence lifetime and Raman spectroscopy.

In 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 Abstract An image comparison between FLIm and Raman spectroscopy acquired with the bimodal probe onseveral tissue samples.

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.

Raman-on-chip device and detection fibres with fibre Bragg grating for analysis of solutions and particles.

An all-fibre based Raman-on-chip setup is introduced which enables analysis of solutions and trapped particles without microscopes or objectives. Beside the novel quartz microfluidic chip, innovative multi-core single-mode fibres with integrated fibre Bragg gratings are used for detection. The limit of quantitation is 7.5 mM for urea and 2.5 mM for nicotine with linear Raman spectroscopy. This is an improvement of more than two orders of magnitude compared with previous fibre-based microfluidic Raman detection schemes. Furthermore, our device was combined with optical traps to collect Raman-on-chip spectra of spherical polymer beads.

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.

Automated classification of healthy and keloidal collagen patterns based on processing of SHG images of human skin.

All-optical microspectroscopic and tomographic tools have a great potential for the clinical investigation of human skin and skin diseases. However, automated optical tomography or even microscopy generate immense data sets. Therefore, in order to implement such diagnostic tools into the medical practice in both hospitals and private practice, there is a need for automated data handling and image analysis ideally implementing automized scores to judge the physiological state of a tissue section. In this contribution, the potential of an image processing algorithm for the automated classification of skin into normal or keloid based on second-harmonic generation (SHG) microscopic images is demonstrated. Such SHG data is routinely recorded within a multimodal imaging approach. The classification of the tissue implemented in the algorithm employs the geometrical features of collagen patterns that differ depending on the constitution, i.e., physiological status of the skin.

Nanoparticle layer deposition for plasmonic tuning of microstructured optical fibers.

Plasmonic nanoparticles with spectral properties in the UV-to-near-IR range have a large potential for the development of innovative optical devices. Similarly, microstructured optical fibers (MOFs) represent a promising platform technology for fully integrated, next-generation plasmonic devices; therefore, the combination of MOFs and plasmonic nanoparticles would open the way for novel applications, especially in sensing applications. In this Full Paper, a cost-effective, innovative nanoparticle layer deposition (NLD) technique is demonstrated for the preparation of well-defined plasmonic layers of selected particles inside the channels of MOFs. This dynamic chemical deposition method utilizes a combination of microfluidics and self-assembled monolayer (SAM) techniques, leading to a longitudinal homogeneous particle density as long as several meters. By using particles with predefined plasmonic properties, such as the resonance wavelength, fibers with particle-adequate spectral characteristics can be prepared. The application of such fibers for refractive-index sensing yields a sensitivity of about 78 nm per refractive index unit (RIU). These novel, plasmonically tuned optical fibers with freely selected, application-tailored optical properties present extensive possibilities for applications in localized surface plasmon resonance (LSPR) sensing.

Multimodal imaging to study the morphochemistry of basal cell carcinoma.

Basal cell carcinoma is the most abundant malignant neoplasm in humans, the pathology of which is characterized by an abnormal proliferation of basal cells. Basal cell carcinoma can show a variety of different morphologies, which are based on different cellular biology. Furthermore, the carcinoma often grows invisibly to the eye imbedded in the surrounding skin. Therefore, in some cases its clinical detection is challenging. Thus, our work aims at establishing an unsupervised tissue classification method based on multimodal imaging and the application of chemometrics to discriminate basal cell carcinoma from non-diseased tissue. A case study applying multimodal imaging to ex-vivo sections of basal cell carcinoma is presented. In doing so, we apply a combination of various linear and non-linear imaging modalities, i.e. fluorescence, Raman and second-harmonic generation microscopy, to study the morphochemistry of basal cell carcinoma. The joint information content obtained by such multimodal approach in studying various aspects of the malignant tissue alterations associated with basal cell carcinoma is discussed.

Single-pulse fiber Bragg gratings and specific coatings for use at elevated temperatures.

The technique of recording fiber Bragg gratings (FBGs) with single exposure pulses during the fiber drawing process allows production of such gratings in complex array structures, with high mechanical strength of the fiber and in a simple and cost-efficient way. This is of special interest for the growing field of fiber sensor applications with FBGs. A general advantage of fiber sensor systems is their ability to be used also at elevated temperatures compared with conventional electric or electronic sensors. For this purpose, the fiber itself as well as the grating structure and the fiber coating should be stable under such elevated temperature conditions. We have investigated different coating materials and possibilities of making temperature-stable FBGs of types I and II in the range of 100 degrees C-1000 degrees C with good reflection efficiency by single-pulse exposure during the fiber drawing process.