Label‐free spectroscopic tissue characterization using fluorescence excitation‐scanning spectral imaging

Favreau, Peter F.; Deal, Joshua; Harris, Bradley; Weber, David S.; Rich, Thomas C.; Leavesley, Silas J.

doi:10.1002/jbio.201900183

Cited by 11 publications

(11 citation statements)

References 47 publications

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“…Hyperspectral imaging. Hyperspectral imaging analysis approaches were used to examine the distribution and expression of tdTomato fluorescent protein across a range of tissue types, while accounting for tissue type-specific autofluorescence (12). Hyperspectral z-stack images were acquired using a Nikon A1R confocal microscope and several objectives, including ϫ4 (Plan Apo ϫ4), ϫ20 (Plan Fluor ϫ20 multi immersion DIC N2), and ϫ60 (Plan Apo VC 60X WI DIC N2).…”

Section: Methodsmentioning

confidence: 99%

Development of an endothelial cell-restricted transgenic reporter rat: a resource for physiological studies of vascular biology

Alexeyev

Geurts

Annamdevula

et al. 2020

American Journal of Physiology-Heart and Circulatory Physiology

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Here, we report the generation of a Cre-recombinase transgenic rat, where iCre is driven using a CDH5 promoter. The CDH5 promoter was cloned from rat pulmonary microvascular endothelial cells and demonstrated approximately 60% similarity to the murine counterpart. The cloned rat promoter was 2,508 bp, it extended 79 bp beyond the transcription start site, and it was 22,923 bp upstream of the translation start site. The novel promoter was cloned upstream of codon-optimized iCre and subcloned into a Sleeping Beauty transposon vector for transpositional transgenesis in Sprague-Dawley rats. Transgenic founders were generated and selected for iCre expression. Crossing the CDH5-iCre rat with a tdTomato reporter rat resulted in progeny displaying endothelial-restricted fluorescence. tdTomato fluorescence was prominent in major arteries and veins, and it was similar in males and females. Quantitative analysis of the carotid artery and the jugular vein revealed that on average more than 50% of the vascular surface area exhibited strong fluorescence. tdTomato fluorescence was observed in the circulations of every tissue tested. The microcirculation in all tissues tested displayed homogenous fluorescence. Fluorescence was examined across young (6-7.5 months), middle (14-16.5 months), and old age (17-19.5 months) groups. Although tdTomato fluorescence was seen in middle- and old-age animals, the intensity of the fluorescence was significantly reduced compared to that seen in the young rats. Thus, this endothelial restricted transgenic rat offers a novel platform to test endothelial microheterogeneity within all vascular segments and it provides exceptional resolution of endothelium within organ microcirculations for application to translational disease models.

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Section: Methodsmentioning

confidence: 99%

Development of an endothelial cell-restricted transgenic reporter rat: a resource for physiological studies of vascular biology

Alexeyev

Geurts

Annamdevula

et al. 2020

American Journal of Physiology-Heart and Circulatory Physiology

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“…The development of optical biopsy for multipurpose diagnostic applications in biomedicine relies on the presence of numerous types of biomolecules acting as endogenous fluorophores (EFs) [1][2][3]. In hepatology, multifaceted diagnostic perspectives are accounted for by the biochemical complexity of the liver and its engagement in numerous metabolic and catabolic functions to maintain the systemic homeostasis of nutrients and micronutrients, such as proteins, lipids, carbohydrates, vitamins, minerals, and electrolytes, and to detoxify the organism [4][5][6].…”

Section: Introductionmentioning

confidence: 99%

Spectrofluorometric Analysis of Autofluorescing Components of Crude Serum from a Rat Liver Model of Ischemia and Reperfusion

et al. 2020

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Autofluorescence (AF) of crude serum was investigated with reference to the potential of its intrinsic AF biomarkers for the noninvasive diagnosis of liver injury. Spectral parameters of pure compounds representing retinol (vitamin A) and fluorescing free fatty acids were characterized by spectrofluorometry, to assess spectral parameters for the subsequent AF analysis of serum, collected from rats undergoing liver ischemia/reperfusion (I/R). Differences in AF spectral profiles detected between control and I/R were due to the increase in the AF components representing fatty acids in I/R serum samples. No significant changes occurred for retinol levels, consistently with the literature reporting that constant retinol levels are commonly observed in the blood, except for malnutrition or chronic severe liver disease. Conversely, fatty acids, in particular arachidonic and linoleic acid and their derivatives, act as modulating agents in inflammation, representing both a protective and damaging response to stress stimuli. The biometabolic and pathophysiological meaning of serum components and the possibility of their direct detection by AF spectrofluorometry open up interesting perspectives for the development of AF serum analysis, as a direct, cost effective, supportive tool to assess liver injury and related systemic metabolic alterations, for applications in experimental biomedicine and foreseen translation to the clinics.

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“…To our knowledge, the comparison of two different DF ELSS HSI detection principles in terms of tissue identification was conducted for the first time. So far, HSI only coupled with other spectral techniques was applied for the differentiation of healthy and cancerous tissue, such as Raman [ 39 ], fluorescence [ 40 ], or FT-IR imaging [ 25 ].…”

Section: Discussionmentioning

confidence: 99%

Comparison of Whiskbroom and Pushbroom darkfield elastic light scattering spectroscopic imaging for head and neck cancer identification in a mouse model

et al. 2021

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The early detection of head and neck cancer is a prolonged challenging task. It requires a precise and accurate identification of tissue alterations as well as a distinct discrimination of cancerous from healthy tissue areas. A novel approach for this purpose uses microspectroscopic techniques with special focus on hyperspectral imaging (HSI) methods. Our proof-of-principle study presents the implementation and application of darkfield elastic light scattering spectroscopy (DF ELSS) as a non-destructive, high-resolution, and fast imaging modality to distinguish lingual healthy from altered tissue regions in a mouse model. The main aspect of our study deals with the comparison of two varying HSI detection principles, which are a point-by-point and line scanning imaging, and whether one might be more appropriate in differentiating several tissue types. Statistical models are formed by deploying a principal component analysis (PCA) with the Bayesian discriminant analysis (DA) on the elastic light scattering (ELS) spectra. Overall accuracy, sensitivity, and precision values of 98% are achieved for both models whereas the overall specificity results in 99%. An additional classification of model-unknown ELS spectra is performed. The predictions are verified with histopathological evaluations of identical HE-stained tissue areas to prove the model’s capability of tissue distinction. In the context of our proof-of-principle study, we assess the Pushbroom PCA-DA model to be more suitable for tissue type differentiations and thus tissue classification. In addition to the HE-examination in head and neck cancer diagnosis, the usage of HSI-based statistical models might be conceivable in a daily clinical routine. Graphical abstract

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Label‐free spectroscopic tissue characterization using fluorescence excitation‐scanning spectral imaging

Cited by 11 publications

References 47 publications

Development of an endothelial cell-restricted transgenic reporter rat: a resource for physiological studies of vascular biology

Development of an endothelial cell-restricted transgenic reporter rat: a resource for physiological studies of vascular biology

Spectrofluorometric Analysis of Autofluorescing Components of Crude Serum from a Rat Liver Model of Ischemia and Reperfusion

Comparison of Whiskbroom and Pushbroom darkfield elastic light scattering spectroscopic imaging for head and neck cancer identification in a mouse model

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