Rapid biodiagnostic ex vivo imaging at 1 μm pixel resolution with thermal source FTIR FPA

Findlay, Catherine Rui Jin; Wiens, Richard; Rak, Margaret; Sedlmair, Julia; Hirschmugl, Carol J.; Morrison, Jason; Mundy, C. J.; Kansiz, Mustafa; Gough, Kathleen

doi:10.1039/c4an01982b

Cited by 48 publications

(56 citation statements)

References 19 publications

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“…20,24 Following the availability of commercial benchtop IR imaging system with an improved optical set-up, high-resolution imaging has been possible where measurements on cells and tissues were carried out (ref. [34][35][36] with promising outcomes. In this study, IR imaging of colon tissues using the highmagnification set-up in combination with multivariate analysis has been carried out.…”

Section: Resultsmentioning

confidence: 99%

High-resolution FTIR imaging of colon tissues for elucidation of individual cellular and histopathological features

Nallala

Lloyd

Shepherd

et al. 2016

Analyst

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a Novel technologies that could complement current histopathology based cancer diagnostic methods are under examination. In this endeavour mid-infrared spectroscopic imaging is a promising candidate that can provide valuable bio-molecular information from unstained cells and tissues in a rapid and a nondestructive manner. With this imaging technique, the biochemical information obtained from smaller areas of the tissues can be of clinical significance and hence the measured pixel size. Until recently it was difficult to obtain spectral data from pixels below around 5 microns square. High NA objectives have been utilised to reduce the ideal diffraction limit, enabling for the first time elucidation of subcellular features.In this context, the ability of high-resolution imaging, obtained using novel high-magnification optics retro-fitted onto a bench top FTIR imaging system, to characterise histopathological features in colonic tissues has been tested. Formalin fixed paraffin embedded colon tissues from three different pathologies were imaged directly using the conventional and the high-magnification imaging set-ups. To circumvent chemical de-paraffinization protocols, an extended multiplicative signal correction (EMSC) based electronic de-paraffinization was carried out on all the infrared images. Multivariate analysis of the highmagnification infrared imaging data showed a detailed information of the histological features of the colon tissue in comparison to conventional imaging. Furthermore, high-magnification imaging has enabled a label-free characterization of the mucin rich goblet cell features in an unprecedented manner.The current study demonstrates the applicability of high-magnification FTIR imaging to characterise complex tissues on a smaller scale that could be of clinical significance.

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Section: Resultsmentioning

confidence: 99%

High-resolution FTIR imaging of colon tissues for elucidation of individual cellular and histopathological features

Nallala

Lloyd

Shepherd

et al. 2016

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“…This trade-off in spatial resolution produces a larger field of view and is clear when comparing similar results from our previous work. 25 Extracted transmission profiles from elements 4, 5 and 6 from group 7 in HM mode show that none of the elements are fully resolved, which is expected based on previous work 26 and the wavelength of infrared used (3.2 µm) even with a projected pixel size of 1.1 µm. However, contrary to expectations, the contrast between the lines for each element imaged with the lens decreases compared to without the lens in HM mode.…”

Section: Assessment Of Spatial Resolution For High-magnification Ftirmentioning

confidence: 60%

“…25 Advances in microscope design such as with the Agilent Cary 620 system use 'high-magnification' (HM) optics, producing a projected pixel size of 1.1 µm -comparable to ATR 26 . However, the real spatial resolution of the system, R, i.e.…”

Section: Introductionmentioning

confidence: 99%

FTIR spectroscopic imaging and mapping with correcting lenses for studies of biological cells and tissues

et al. 2016

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Histopathology of tissue samples is used to determine the progression of cancer usually by staining and visual analysis. It is recognised that disease progression from healthy tissue to cancerous is accompanied by spectral signature changes in the infrared range. In this work, FTIR spectroscopic imaging in transmission using a focal plane array (96 x 96 pixels) has been applied to the characterisation of Barrett's esophageal adenocarcinoma. Spectral images were acquired in transmission mode. To correct optical aberrations, infrared transparent lenses were used of the same material as the slide on which biopsies were fixed. The lenses acted as an immersion objective, reducing scattering and improving spatial resolution.A novel mapping approach is presented where spectral images obtained with added lens are stitched together such that the dataset contained a representative section of the esophageal tissue. Images were also acquired in transmission mode using high-magnification optics for enhanced spatial resolution, and were also acquired using a germanium micro-ATR objective for comparison. The reduction of scattering was assessed using k-means clustering. The same tissue section map, which contained a region of high grade dysplasia was analysed using hierarchical clustering analysis. A reduction of the trough at 1077 cm -1 in the second derivative spectra was identified as an indicator of high grade dysplasia.In addition, the spatial resolution obtained with the added lens using high-magnification optics was assessed by measurements of a sharp interface of polymer laminate, which was also compared with the spatial resolution achieved in micro ATR-FTIR imaging. The resolution was determined from the change in absorbance of spectral bands from one side of the polymer interface to the other. In transmission mode using the lens, it was determined to be 12 µm and using micro-ATR, the resolution was 3 µm for the band at ca. 6 µm The spatial resolution was also assessed with and without the added lens, in normal and high-magnification mode using a USAF target.Spectroscopic images of cells in transmission using two lenses are also presented, which are necessary for correcting chromatic aberration and refraction in both the condenser and objective. The use of lenses is shown to be necessary in obtaining high-quality spectroscopic images of cells in transmission mode and proves the applicability of the pseudo hemisphere approach for this and other microfluidic systems.2

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“…While synchrotron-FT-IR analyses require dedicated facilities, advances in the development of high-magnification optics have made it possible to perform FPA-based infrared imaging at a spatial resolution comparable with Raman instruments, even without access to a synchrotron beamline (Findlay et al, 2015). Analyses of cells in aqueous suspensions are additionally possible using attenuated total reflectance (ATR)-FT-IR imaging (Kuimova et al, 2009).…”

Section: Ft-ir Imagingmentioning

confidence: 99%

Vibrational Spectroscopy for Imaging Single Microbial Cells in Complex Biological Samples

Harrison

Berry

2017

Front. Microbiol.

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Vibrational spectroscopy is increasingly used for the rapid and non-destructive imaging of environmental and medical samples. Both Raman and Fourier-transform infrared (FT-IR) imaging have been applied to obtain detailed information on the chemical composition of biological materials, ranging from single microbial cells to tissues. Due to its compatibility with methods such as stable isotope labeling for the monitoring of cellular activities, vibrational spectroscopy also holds considerable power as a tool in microbial ecology. Chemical imaging of undisturbed biological systems (such as live cells in their native habitats) presents unique challenges due to the physical and chemical complexity of the samples, potential for spectral interference, and frequent need for real-time measurements. This Mini Review provides a critical synthesis of recent applications of Raman and FT-IR spectroscopy for characterizing complex biological samples, with a focus on developments in single-cell imaging. We also discuss how new spectroscopic methods could be used to overcome current limitations of single-cell analyses. Given the inherent complementarity of Raman and FT-IR spectroscopic methods, we discuss how combining these approaches could enable us to obtain new insights into biological activities either in situ or under conditions that simulate selected properties of the natural environment.

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Rapid biodiagnostic ex vivo imaging at 1 μm pixel resolution with thermal source FTIR FPA

Cited by 48 publications

References 19 publications

High-resolution FTIR imaging of colon tissues for elucidation of individual cellular and histopathological features

High-resolution FTIR imaging of colon tissues for elucidation of individual cellular and histopathological features

FTIR spectroscopic imaging and mapping with correcting lenses for studies of biological cells and tissues

Vibrational Spectroscopy for Imaging Single Microbial Cells in Complex Biological Samples

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