Osteoarthritis (OA), a degenerative joint disease presenting as loss of cartilage, is a leading cause of disability worldwide, increasingly with aging populations. Early detection is crucial for effective treatment since there is no definitive cure, yet, current assessment techniques fall short and rely on ionising radiation or invasive procedures. We report an application of multimodal “spectromics”, low-level abstraction data fusion of non-destructive NIR Raman and NIR-SWIR absorption spectroscopy, providing an enhanced, interpretable “fingerprint” for diagnosis of OA in human cartilage. Under multivariate statistical analyses and supervised machine learning, cartilage was classified with high precision and disease state predicted accurately. Discriminatory features within the spectromics fingerprint elucidated clinically relevant tissue components (OA biomarkers). Further, we have developed an automated goniometric 3D hyperspectral mapping setup, and characterised OA cartilage on whole human femoral heads post hip arthroplasty for spatially-resolved spectromics. These results lay foundation for minimally invasive, deeply penetrating, label-free, chemometric diagnosis of the hip.
Multiphoton microscopies are an invaluable tool in biomedical imaging given their inherent capabilities for label free imaging, optical sectioning, chemical and structural specificity. They comprise various types of Coherent Raman microscopies (CR), such as Coherent Anti-Stokes Raman Scattering (CARS), Stimulated Raman Loss (SRL) or Stimulated Raman Gain, different kinds of Harmonic Generation imaging (HG) such as Second and Third Harmonic Generation (SHG and THG respectively), and Multiphoton Autofluorescence imaging (MA) such as Two and Three Photon Excited Autofluorescence (TPEAF and ThPEAF respectively). Despite their significant advantages, multiphoton microscopies, comparably to all other types of optical microscopies, exhibit limited penetration depth in tissue due to absorption and scattering. In this work we explore the advantages of multiphoton microscopies in hard and soft deep tissue imaging when using excitation wavelengths in the range of Short-Wavelength Infrared (SWIR) windows which occur between 1000 nm and 2500 nm. These spectral windows have notable merits including longer attenuation lengths and none or very low signal absorption observed for almost all kinds of multiphoton microscopy. We show results of using excitations in the SWIR windows, generated by standard as well as novel sources, such as a thulium fibre laser, in different types of multiphoton microscopy on a variety of hard and soft tissue samples (bone, cartilage and other tissue types) and demonstrate the advantages of using excitations in this wavelength range, including longer penetration depth and high resolution for deep tissue imaging.
Multiphoton imaging methods such as Coherent Raman Scattering (CRS) microscopy which also comprises Second
Harmonic Generation (SHG) and Two Photon Excited Auto-Fluorescence (TPEAF) imaging (termed as multimodal
Coherent Raman microscopy), have greatly facilitated the advancement of biomedical research due to their unique
features. Multimodal CRS microscopy, is label free, chemically specific, inherently ‘confocal’ offering three independent
contrast mechanisms which can be associated in a composite image comprising a wide range of chemical and structural
information about the interrogated sample. The standard light source for multimodal CRS microscopy is a picosecond
pumped Optical Parametric Oscillator (OPO) which has exhibited excellent performance but due to its associated high
cost, maintenance, complexity and requirement of a dedicated optics laboratory, has hindered the wider adoption of
multimodal CRS microscopy and especially its deployment in clinical applications.
Here we present a novel, low cost Optical Parametric Amplifier (OPA) based on a MgO doped Periodically Poled Lithium
Niobate (PPLN) crystal seeded by a continuous wave (CW) laser source and pumped by a picosecond laser at 1031nm,
which removes any synchronisation requirements. We show that this OPA is a versatile light source module that can be
tailored to the tunability and affordability requirements of the specific application. We demonstrate that it can be used
either in association with an OPO or on its own as a light source for multimodal CRS microscopy and we show its
performance by imaging a variety of standards and biological samples.
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