Two-color widefield fluorescence microendoscopy enables multiplexed molecular imaging in the alveolar space of human lung tissue

Abstract

Citation for published item:urstji¡ D xikol nd ekrmD ehsn F nd ghoudhryD ushr F nd whonldD xeil nd nnerD wihel qF nd edrettiD ittore nd hlgrnoD ul eF nd hole(eldD imm nd qirkinD tohn wF nd wooreD enne nd frdleyD wrk nd hhliwlD uevin @PHITA 9woEolor wide(eld )uoresene miroendosopy enles multiplexed moleulr imging in the lveolr spe of humn lung tissueF9D tournl of iomedil optisFD PI @RAF HRTHHWF Further information on publisher's website: Use policyThe full-text may be used and/or reproduced, and given to third parties in any format or medium, without prior permission or charge, for personal research or study, educational, or not-for-prot purposes provided that:• a full bibliographic reference is made to the original source • a link is made to the metadata record in DRO • the full-text is not changed in any way The full-text must not be sold in any format or medium without the formal permission of the copyright holders.Please consult the full DRO policy for further details. Abstract. We demonstrate a fast two-color widefield fluorescence microendoscopy system capable of simultaneously detecting several disease targets in intact human ex vivo lung tissue. We characterize the system for light throughput from the excitation light emitting diodes, fluorescence collection efficiency, and chromatic focal shifts. We demonstrate the effectiveness of the instrument by imaging bacteria (Pseudomonas aeruginosa) in ex vivo human lung tissue. We describe a mechanism of bacterial detection through the fiber bundle that uses blinking effects of bacteria as they move in front of the fiber core providing detection of objects smaller than the fiber core and cladding (∼3 μm). This effectively increases the measured spatial resolution of 4 μm. We show simultaneous imaging of neutrophils, monocytes, and fungus (Aspergillus fumigatus) in ex vivo human lung tissue. The instrument has 10 nM and 50 nM sensitivity for fluorescein and Cy5 solutions, respectively. Lung tissue autofluorescence remains visible at up to 200 fps camera acquisition rate. The optical system lends itself to clinical translation due to high-fluorescence sensitivity, simplicity, and the ability to multiplex several pathological molecular imaging targets simultaneously.

Image computing for fibre-bundle endomicroscopy: A review

“…The probes' miniature sizes enable use through the working channel of most commercially available endoscopes as well as some needles/catheters, while the relevant high data acquisition rate (>12fps) enables realtime imaging of moving structures, resulting in Cellvizio (pCLE) being the most widely used endomicroscopy platform approved for clinical use. A rapidly growing volume of alternative, non-commercial experimental architectures is being developed, including (i) right-angle stage attachment for standard desktop confocal microscopes (ii) line-scanning confocal endomicroscopy Yang, 2015, 2016), improving the acquisition frame rate without compromising substantially image quality; (iii) flexible and low-cost endomicroscopy architectures (Hong et al, 2016;Krstajić et al, 2016;Pierce et al, 2011;Shin et al, 2010), employing LED, widefield illumination; (iv) structured illumination endomicroscopy (Bozinovic et al, 2008;Ford et al, 2012b;Ford and Mertz, 2013), providing out-of-focus background rejection without beam scanning;…”

“…Examples illustrating properties of coherent fibre bundles that limit the imaging capabilities of fibred endoscopy. (a) Scanning Electron Microscopy (SEM) image of commercial coherent fibre bundle (FIGH-30-650S, Fujikura), along with (b) a uniform, flood illumination (520nm) image of the same fibre bundle, using widefield endomicroscopy (Krstajić et al, 2016). The variable size and shape, as well as irregular distribution of the cores is apparent in both (a) and (b).…”

Development of an Alveolar Transbronchial Catheter for Concurrent Fiber Optics-Based Imaging and Fluid Delivery

“…For potential methods of identifying infection, several teams have developed imaging markers used with pCLE to identify bacterial infections [17] both systemically and in situ [18], 19]. Of particular interest is a two-color methodology that has shown the potential to identify neutrophil recruitment, fungal infection, and bacterial presence in ex vivo human lungs [20].…”

Patch-Based Sparse Representation For Bacterial Detection

“…In this work, we investigate the performance of an unsupervised approach for bacterial detection in datasets of optical endomicroscopy (OEM) lung images [15][16][17][18][19][20][21]. The main contributions of this work are threefold.…”