Xia FTIRI Lab
We have a PerkinElmer Spotlight 300 imager, which consists of a classical FTIR spectrophotometer coupled to an infrared microscope. The pixel size of the liquid-nitrogen-cooled detector is 6.25 µm. We also have an ATR attachment, which can improve the pixel resolution in this system down to 1.56µm.
We have published so far 16 journal papers on the studies of cartilage and its degradation by FTIRI.
Y Xia, N Ramakrishnan, A Bidthanapally, “The Depth-dependent Anisotropy of Articular Cartilage by Fourier-Transform Infrared Imaging (FTIRI)”, Osteoarthritis and Cartilage, 15 (7), 780-788 (2007).
What are the orientational anisotropies of different chemical components in cartilage? Although FTIRI does not have the resolution to identify individual collagen fibrils, we show for the first time that a new type of quantitative IR imaging (the absorbance anisotropy map) can reveal the angular-dependent variations of the chemicals in cartilage by eliminating other biochemical and instrumental factors contributing to the variation in infrared absorbance images.
N Ramakrishnan, Y Xia, A Bidthanapally, “Polarized IR Microscopic Imaging of Articular Cartilage”, Physics in Medicine and Biology, 52 (15), 4601-4614 (2007).
What happens if you polarize the infrared light in imaging? This report discusses the infrared absorption anisotropy of articular cartilage under polarized infrared radiation as well as cross-polarization.
N Ramakrishnan, Y Xia, A Bidthanapally, M Lu, “Determination of Zonal Boundaries in Articular Cartilage Using Infrared Dichroism”, Applied Spectroscopy, 61 (12), 1404-1409 (2007).
Can we sub-divide the thickness of articular cartilage into structural zones according to its chemical information? A new method was developed for the first time in FTIRI, which uses infrared dichroism of the amide components to determine the zonal boundaries in articular cartilage. This method is independent of many instrumental and specimen variations, and hence practical and robust.
Y Xia, H Alhadlaq, N Ramakrishnan, A Bidthanapally, F Badar, M Lu, “Molecular and Morphological Adaptations in Compressed Articular Cartilage by Polarized Light Microscopy and Fourier-Transform Infrared Imaging”, Journal of Structural Biology, 164 (1), 88-95 (2008).
What are the changes in molecular bond directions when we image cartilage while it is being compressed (since it is a load-bearing tissue)? An excellent correlation is found between the relative depth of the minimum retardance in PLM and the relative depth of the Amide II anisotropic cross-over in FTIRI. The changes in amide anisotropies in different deformed histological zones are explained by a model that describes the strain-dependent tipping angle of the amide bonds. Our images appeared on the cover of this issue.
N Ramakrishnan, Y Xia, A Bidthanapally, “Fourier-transform infrared anisotropy in cross and parallel sections of tendon and articular cartilage”, Journal of Orthopaedic Surgery and Research, 3 (1), 48 (2008).
Are there differences if cartilage is imaged from three orthogonal directions? The infrared anisotropy of tendon as well as cartilage has been investigated in regular, parallel and cross sections from a 3D tissue block. An interesting situation occurs when the long axis of the fibrils is perpendicular to the plane of the tissue section. Though the infrared anisotropy of amide components in tendon cross sections would be expected to be isotropic, the experimental results show a clear anisotropy for both amide I and amide II components in tendon. This could be attributed to the zigzag nature of the collagen fibers in tendon.
JH Yin, Y Xia, “Macromolecular Concentrations in Bovine Nasal Cartilage by Fourier Transform Infrared Imaging and Principal Component Regression”, Applied Spectroscopy, 64 (11), 1199-1208 (2010).
How can we quantify the major molecular components in cartilage in infrared imaging? This combined FTIRI-PCR and biochemistry project successfully determined the concentration distributions of principal molecular components (both collagen and proteoglycan) in bovine nasal cartilage quantitatively and accurately.
Y Xia, D Mittelstaedt, N Ramakrishnan, M Szarko, A Bidthanapally, “Depth-Dependent Anisotropies of Amides and Sugar in Perpendicular and Parallel Sections of Articular Cartilage by Fourier Transform Infrared Imaging (FTIRI)”, Microscopy Research and Technique, 74 (2), 122-132 (2011).
What happens if you look directly at the ends of the collagen fibrils? (just like looking directly at the head of an arrow) This is the first infrared investigation of the dipolar bond anisotropies of amides and sugar in articular cartilage over the entire depth of the non-calcified cartilage, based on the imaging results of parallel tissue sections.
JH Yin, Y Xia, “Chemical Visualization of Individual Chondrocytes in Articular Cartilage by Attenuated-Total-Reflection Fourier Transform Infrared Microimaging”, Biomedical Optics Express, 2 (4), 937-945 (2011). (PMC3072132)
Can you visualize the chemical structure in/around a cell? This is the first chemical visualization of the territorial matrix of fine collagen fibrils surrounding the individual chondrocytes by high resolution ATR-FTIRM at 1.56µm. The polarization experiments used the absorption ratio of amide I to amide II bands.
JH Yin, Y Xia, N Ramakrishnan, “Depth-dependent Anisotropy of Proteoglycan in Articular Cartilage by Fourier Transform Infrared Imaging”, Vibrational Spectroscopy, 57 (2), 338-341 (2011). (PMID: 22025814; PMCID: PMC3197785)
Is sugar in cartilage isotropic by the infrared measurement? An anisotropic flipping point of the sugar band in the infrared experiment was noticed for the first time, by the FTIRI measurement and PCR calculation. This flipping point is not near the surface but at the deepest part of the radial zone approaching the tidemark. (In comparison, an anisotropic flipping point of the amides was founded by our early papers in 2007 (the 1st paper above). The amide flopping point is near the surface, around the center of the transitional zone.)
JH Yin, Y Xia, M Lu, “Concentration Profiles of Collagen and Proteoglycan in Articular Cartilage by Fourier Transform Infrared Imaging and Principal Component Regression”, Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy, 88, 90-96 (2012). (PMID: 22197357; PMCID: PMC3268907)
Mapping the concentration of both collagen and proteoglycan in articular cartilage. The simultaneous construction of both collagen and proteoglycan concentration images demonstrates that this combined imaging and chemometrics approach could be used as a sensitive tool to accurately resolve and visualize the concentration distributions of macromolecules in biological tissues.