Large scale infrared imaging of tissue micro arrays (TMAs) using a tunable Quantum Cascade Laser (QCL) based microscope

Bassan, Paul; Weida, Miles J.; Rowlette, Jeremy; Gardner, Peter

doi:10.1039/c4an00638k

Cited by 116 publications

(83 citation statements)

References 15 publications

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“…Nevertheless, we are currently developing FTIR acquisition with new devices, such as infrared focal plane arrays, making it possible to reduce acquisition time and consequently, increase the spatial resolution. 26 It has been reported that visual assessment of IF with scoring (IFTA) according to the Banff classification suffers from poor interobserver reproducibility. 3,27 Moreover, the use of a small number of grades to describe the severity of individual histologic injuries may lack sensitivity during the early stage of IFTA.…”

Section: Discussionmentioning

confidence: 99%

Renal Graft Fibrosis and Inflammation Quantification by an Automated Fourier–Transform Infrared Imaging Technique

Vuiblet

Fere

Gobinet

et al. 2015

JASN

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Renal interstitial fibrosis and interstitial active inflammation are the main histologic features of renal allograft biopsy specimens. Fibrosis is currently assessed by semiquantitative subjective analysis, and color image analysis has been developed to improve the reliability and repeatability of this evaluation. However, these techniques fail to distinguish fibrosis from constitutive collagen or active inflammation. We developed an automatic, reproducible Fourier-transform infrared (FTIR) imaging-based technique for simultaneous quantification of fibrosis and inflammation in renal allograft biopsy specimens. We generated and validated a classification model using 49 renal biopsy specimens and subsequently tested the robustness of this classification algorithm on 166 renal grafts. Finally, we explored the clinical relevance of fibrosis quantification using FTIR imaging by comparing results with renal function at 3 months after transplantation (M3) and the variation of renal function between M3 and M12. We showed excellent robustness for fibrosis and inflammation classification, with .90% of renal biopsy specimens adequately classified by FTIR imaging. Finally, fibrosis quantification by FTIR imaging correlated with renal function at M3, and the variation in fibrosis between M3 and M12 correlated well with the variation in renal function over the same period. This study shows that FTIRbased analysis of renal graft biopsy specimens is a reproducible and reliable label-free technique for quantifying fibrosis and active inflammation. This technique seems to be more relevant than digital image analysis and promising for both research studies and routine clinical practice.

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Section: Discussionmentioning

confidence: 99%

Renal Graft Fibrosis and Inflammation Quantification by an Automated Fourier–Transform Infrared Imaging Technique

Vuiblet

Fere

Gobinet

et al. 2015

JASN

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“…If the information content is highly localized in the spectral domain, large increases in throughput can be achieved. Bassan et al showed that a Spero Ò microscope is able to collect single-wavenumber images of a breast tissue microarray that contains more than 26 million spectra in 540 seconds, compared with more than 17 hours using an FTIR microscope (15). This allowed an entire biopsy sample covering an area of 20£24 mm to be imaged with 4 micron pixel size in 9 minutes.…”

Section: Introductionmentioning

confidence: 99%

Sensitivity Advantage of QCL Tunable-Laser Mid-Infrared Spectroscopy Over FTIR Spectroscopy

Childs

Hogg

Revin

et al. 2015

Applied Spectroscopy Reviews

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Interest in mid-infrared spectroscopy instrumentation beyond classical FTIR using a thermal light source has increased dramatically in recent years. Synchrotron, supercontinuum, and external-cavity quantum cascade laser light sources are emerging as viable alternatives to the traditional thermal black-body emitter (Globar), especially for remote interrogation of samples ("stand-off" detection) and for hyperspectral imaging at diffraction-limited spatial resolution ("microspectroscopy"). It is thus timely to rigorously consider the relative merits of these different light sources for such applications. We study the theoretical maximum achievable signal-to-noise ratio (SNR) of FTIR using synchrotron or supercontinuum light vs. that of a tunable quantum cascade laser, by reinterpreting an important result that is well known in near-infrared optical coherence tomography imaging. We rigorously show that mid-infrared spectra can be acquired up to 1000 times fasterusing the same detected light intensity, the same detector noise level, and without loss of SNR-using the tunable quantum cascade laser as compared with the FTIR approach using synchrotron or supercontinuum light. We experimentally demonstrate the effect using a novel, rapidly tunable quantum cascade laser that acquires spectra at rates of up to 400 per second. We also estimate the maximum potential spectral acquisition rate of our prototype system to be 100,000 per second.

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“…Correlative IR/SMLM studies are expected to become significantly more approachable with new benchtop light-sources that are brighter than traditional Globars and thus deliver better signal-to-noise faster allowing for nonsynchrotron live-cell experiments 52,53 . Furthermore, robust water absorption correction 54 will allow for examination of the Amide I band and better elucidation of changes to protein composition.…”

Section: Discussionmentioning

confidence: 99%

Correlative Synchrotron Fourier Transform Infrared Spectroscopy and Single Molecule Super Resolution Microscopy for the Detection of Composition and Ultrastructure Alterations in Single Cells

Whelan

Bell

2015

ACS Chem. Biol.

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Single molecule localization microscopy (SMLM) and synchrotron Fourier transform infrared (S-FTIR) spectroscopy are two techniques capable of elucidating unique and valuable biological detail. SMLM provides images of the structures and distributions of targeted biomolecules at spatial resolutions up to an order of magnitude better than the diffraction limit, whereas IR spectroscopy objectively measures the holistic biochemistry of an entire sample, thereby revealing any variations in overall composition. Both tools are currently applied extensively to detect cellular response to disease, chemical treatment, and environmental change. Here, these two techniques have been applied correlatively at the single cell level to probe the biochemistry of common fixation methods and have detected various fixation-induced losses of biomolecular composition and cellular ultrastructure. Furthermore, by extensive honing and optimizing of fixation protocols, many fixation artifacts previously considered pervasive and regularly identified using IR spectroscopy and fluorescence techniques have been avoided. Both paraformaldehyde and two-step glutaraldehyde fixation were identified as best preserving biochemistry for both SMLM and IR studies while other glutaraldehyde and methanol fixation protocols were demonstrated to cause significant biochemical changes and higher variability between samples. Moreover, the potential complementarity of the two techniques was strikingly demonstrated in the correlated detection of biochemical changes as well as in the detection of fixation-induced damage that was only revealed by one of the two techniques.

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Large scale infrared imaging of tissue micro arrays (TMAs) using a tunable Quantum Cascade Laser (QCL) based microscope

Cited by 116 publications

References 15 publications

Renal Graft Fibrosis and Inflammation Quantification by an Automated Fourier–Transform Infrared Imaging Technique

Renal Graft Fibrosis and Inflammation Quantification by an Automated Fourier–Transform Infrared Imaging Technique

Sensitivity Advantage of QCL Tunable-Laser Mid-Infrared Spectroscopy Over FTIR Spectroscopy

Correlative Synchrotron Fourier Transform Infrared Spectroscopy and Single Molecule Super Resolution Microscopy for the Detection of Composition and Ultrastructure Alterations in Single Cells

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