scholarly journals Reliable Assessment and Quantification of the Fluorescence-Labeled Antisense OligonucleotidesIn Vivo

2014 ◽  
Vol 2014 ◽  
pp. 1-10
Author(s):  
Maria Chiara Munisso ◽  
Tetsuji Yamaoka
Keyword(s):  
Fluorescence Intensity ◽  
Near Infrared ◽  
Fluorescent Dyes ◽  
Large Change ◽  
Optical Systems ◽  
Stable Complex ◽  
Tissue Samples ◽  
Reliable Assessment ◽  
New Methodologies

The availability of fluorescent dyes and the advances in the optical systems forin vivoimaging have stimulated an increasing interest in developing new methodologies to study and quantify the biodistribution of labeled agents. However, despite these great achievements, we are facing significant challenges in determining if the observed fluorescence does correspond to the quantity of the dye in the tissues. In fact, although the far-red and near-infrared lights can propagate through several centimetres of tissue, they diffuse within a few millimetres as consequence of the elastic scattering of photons. In addition, when dye-labeled oligonucleotides form stable complex with cationic carriers, a large change in the fluorescence intensity of the dye is observed. Therefore, the measured fluorescence intensity is altered by the tissue heterogeneity and by the fluctuation of dye intensity. Hence, in this study a quantification strategy for fluorescence-labeled oligonucleotides was developed to solve these disadvantageous effects. Our results proved that upon efficient homogenization and dilution with chaotropic agents, such as guanidinium thiocyanate, it is possible to achieve a complete fluorescence intensity recovery. Furthermore, we demonstrated that this method has the advantage of good sensitivity and reproducibility, as well as easy handling of the tissue samples.

scholarly journals The Use of Fluorescent Anti-CEA Antibodies to Label, Resect and Treat Cancers: A Review

Biomolecules ◽  
2021 ◽  
Vol 11 (12) ◽  
pp. 1819
Author(s):  
Michael A. Turner ◽  
Thinzar M. Lwin ◽  
Siamak Amirfakhri ◽  
Hiroto Nishino ◽  
Robert M. Hoffman ◽  
...  
Keyword(s):  
Near Infrared ◽  
Fluorescent Dyes ◽  
Review Article ◽  
In Vivo Studies ◽  
Murine Models ◽  
Exclusion Criteria ◽  
Major Barrier ◽  
Tumor Margins ◽  
Cancer Types

A major barrier to the diagnosis and effective treatment of solid-tumor cancers is the difficulty in detection and visualization of tumor margins in primary and metastatic disease. The use of fluorescence can augment the surgeon’s ability to detect cancer and aid in its resection. Several cancer types express carcinoembryonic antigen (CEA) including colorectal, pancreatic and gastric cancer. Antibodies to CEA have been developed and tagged with near-infrared fluorescent dyes. This review article surveyed the use of CEA antibodies conjugated to fluorescent probes for in vivo studies since 1990. PubMed and Google Scholar databases were queried, and 900 titles and abstracts were screened. Fifty-nine entries were identified as possibly meeting inclusion/exclusion criteria and were reviewed in full. Forty articles were included in the review and their citations were screened for additional entries. A total of 44 articles were included in the final review. The use of fluorescent anti-CEA antibodies has been shown to improve detection and resection of tumors in both murine models and clinically. The cumulative results indicate that fluorescent-conjugated anti-CEA antibodies have important potential to improve cancer diagnosis and surgery. In an emerging technology, anti-CEA fluorescent antibodies have also been successfully used for photoimmunotherapy treatment for cancer.

scholarly journals In Vivo Resolution of Multiexponential Decays of Multiple Near-Infrared Molecular Probes by Fluorescence Lifetime-Gated Whole-Body Time-Resolved Diffuse Optical Imaging

2007 ◽  
Vol 6 (4) ◽  
pp. 7290.2007.00020 ◽  
Author(s):  
Walter Akers ◽  
Frederic Lesage ◽  
Dewey Holten ◽  
Samuel Achilefu
Keyword(s):  
Optical Imaging ◽  
Fluorescence Lifetime ◽  
Fluorescence Intensity ◽  
Near Infrared ◽  
Molecular Probes ◽  
Whole Body ◽  
Diffuse Optical Imaging ◽  
Fluorescence Lifetimes ◽  
Time Resolved

The biodistribution of two near-infrared fluorescent agents was assessed in vivo by time-resolved diffuse optical imaging. Bacteriochlorophyll a (BC) and cypate-glysine-arginine-aspartic acid-serine-proline-lysine-OH (Cyp-GRD) were administered separately or combined to mice with subcutaneous xenografts of human breast adenocarcinoma and slow-release estradiol pellets for improved tumor growth. The same excitation (780 nm) and emission (830 nm) wavelengths were used to image the distinct fluorescence lifetime distribution of the fluorescent molecular probes in the mouse cancer model. Fluorescence intensity and lifetime maps were reconstructed after raster-scanning whole-body regions of interest by time-correlated single-photon counting. Each captured temporal point-spread function (TPSF) was deconvolved using both a single and a multiexponental decay model to best determine the measured fluorescence lifetimes. The relative signal from each fluorophore was estimated for any region of interest included in the scanned area. Deconvolution of the individual TPSFs from whole-body fluorescence intensity scans provided corresponding lifetime images for comparing individual component biodistribution. In vivo fluorescence lifetimes were determined to be 0.8 ns (Cyp-GRD) and 2 ns (BC). This study demonstrates that the relative biodistribution of individual fluorophores with similar spectral characteristics can be compartmentalized by using the time-domain fluorescence lifetime gating method.

Small molecular interaction-based fluorescence enhancement for second near-infrared imaging

Nanomedicine ◽  
2020 ◽  
Vol 15 (2) ◽  
pp. 115-129 ◽  
Author(s):  
Wang Zian ◽  
Liu Yang ◽  
Wang Peng ◽  
Jiang Yifei ◽  
Ji Min
Keyword(s):  
Indocyanine Green ◽  
Fluorescence Intensity ◽  
Low Dose ◽  
Near Infrared ◽  
Fluorescence Enhancement ◽  
Infrared Imaging ◽  
Cationic Liposome ◽  
High Temporal Resolution ◽  
New Strategy

Aim: This study described a new strategy to enhance second near-infrared (NIR-II) fluorescence intensity. Materials & methods: NIR-II liposomes were prepared by thin film hydration method and their fluorescence properties were evaluated. The efficacy of the optimized liposome was then evaluated in vivo with low dose and irradiation. Results: Indocyanine green-IR1061 liposome exhibited higher fluorescence intensity (∼fourfold than IR1061 liposome) with the red-shifted emission. The intensity of indocyanine green-IR1061 cationic liposome was enhanced to approximately tenfold, which allowed us to perform angiography with lower doses and less exposure time. Conclusion: We report a new and efficient way to enhance NIR-II fluorescence intensity. This could be used to acquire high temporal resolution and signal-to-background ratio fluorescence imaging.

scholarly journals Comparison of Folate Receptor Targeted Optical Contrast Agents for Intraoperative Molecular Imaging

2015 ◽  
Vol 2015 ◽  
pp. 1-10 ◽  
Author(s):  
Elizabeth De Jesus ◽  
Jane J. Keating ◽  
Sumith A. Kularatne ◽  
Jack Jiang ◽  
Ryan Judy ◽  
...  
Keyword(s):  
Contrast Agents ◽  
Animal Studies ◽  
Near Infrared ◽  
Fluorescent Dyes ◽  
Folate Receptor ◽  
Intraoperative Imaging ◽  
Nir Dyes ◽  
Optical Contrast Agents

Background. Intraoperative imaging can identify cancer cells in order to improve resection; thus fluorescent contrast agents have emerged. Our objective was to do a preclinical comparison of two fluorescent dyes, EC17 and OTL38, which both target folate receptor but have different fluorochromes. Materials. HeLa and KB cells lines were used for in vitro and in vivo comparisons of EC17 and OTL38 brightness, sensitivity, pharmacokinetics, and biodistribution. In vivo experiments were then performed in mice. Results. The peak excitation and emission wavelengths of EC17 and OTL38 were 470/520 nm and 774/794 nm, respectively. In vitro, OTL38 required increased incubation time compared to EC17 for maximum fluorescence; however, peak signal-to-background ratio (SBR) was 1.4-fold higher compared to EC17 within 60 minutes (p<0.001). Additionally, the SBR for detecting smaller quantity of cells was improved with OTL38. In vivo, the mean improvement in SBR of tumors visualized using OTL38 compared to EC17 was 3.3 fold (range 1.48–5.43). Neither dye caused noticeable toxicity in animal studies. Conclusions. In preclinical testing, OTL38 appears to have superior sensitivity and brightness compared to EC17. This coincides with the accepted belief that near infrared (NIR) dyes tend to have less autofluorescence and scattering issues than visible wavelength fluorochromes.

scholarly journals Tracking macrophages in diabetic neuropathy with two-color nanoemulsions for near-infrared fluorescent imaging and microscopy

2021 ◽  
Vol 18 (1) ◽  
Author(s):  
James M. Nichols ◽  
Caitlin V. Crelli ◽  
Lu Liu ◽  
Hoang Vu Pham ◽  
Jelena M. Janjic ◽  
...  
Keyword(s):  
Peripheral Neuropathy ◽  
Fluorescence Microscopy ◽  
Diabetic Neuropathy ◽  
Colloidal Stability ◽  
Near Infrared ◽  
Fluorescent Dyes ◽  
Wild Type ◽  
Nirf Imaging

Abstract Background The incidence of diabetes and diabetic peripheral neuropathy continues to rise, and studies have shown that macrophages play an important role in their pathogenesis. To date, macrophage tracking has largely been achieved using genetically-encoded fluorescent proteins. Here we present a novel two-color fluorescently labeled perfluorocarbon nanoemulsion (PFC-NE) designed to monitor phagocytic macrophages in diabetic neuropathy in vitro and in vivo using non-invasive near-infrared fluorescent (NIRF) imaging and fluorescence microscopy. Methods Presented PFC-NEs were formulated with perfluorocarbon oil surrounded by hydrocarbon shell carrying two fluorescent dyes and stabilized with non-ionic surfactants. In vitro assessment of nanoemulsions was performed by measuring fluorescent signal stability, colloidal stability, and macrophage uptake and subsequent viability. The two-color PFC-NE was administered to Leprdb/db and wild-type mice by tail vein injection, and in vivo tracking of the nanoemulsion was performed using both NIRF imaging and confocal microscopy to assess its biodistribution within phagocytic macrophages along the peripheral sensory apparatus of the hindlimb. Results In vitro experiments show two-color PFC-NE demonstrated high fluorescent and colloidal stability, and that it was readily incorporated into RAW 264.7 macrophages. In vivo tracking revealed distribution of the two-color nanoemulsion to macrophages within most tissues of Leprdb/db and wild-type mice which persisted for several weeks, however it did not cross the blood brain barrier. Reduced fluorescence was seen in sciatic nerves of both Leprdb/db and wild-type mice, implying that the nanoemulsion may also have difficulty crossing an intact blood nerve barrier. Additionally, distribution of the nanoemulsion in Leprdb/db mice was reduced in several tissues as compared to wild-type mice. This reduction in biodistribution appears to be caused by the increased number of adipose tissue macrophages in Leprdb/db mice. Conclusions The nanoemulsion in this study has the ability to identify phagocytic macrophages in the Leprdb/db model using both NIRF imaging and fluorescence microscopy. Presented nanoemulsions have the potential for carrying lipophilic drugs and/or fluorescent dyes, and target inflammatory macrophages in diabetes. Therefore, we foresee these agents becoming a useful tool in both imaging inflammation and providing potential treatment in diabetic peripheral neuropathy.

scholarly journals Optimization of Near-Infrared Fluorescence Voltage-Sensitive Dye Imaging for Neuronal Activity Monitoring in the Rodent Brain

2021 ◽  
Vol 15 ◽  
Author(s):  
Rebecca W. Pak ◽  
Jeeun Kang ◽  
Emad Boctor ◽  
Jin U. Kang
Keyword(s):  
Near Infrared ◽  
Ex Vivo ◽  
Optical Systems ◽  
Clinical Tool ◽  
Voltage Sensitive Dye ◽  
Near Ir ◽  
Indirect Measures ◽  
External Stimuli

Many currently employed clinical brain functional imaging technologies rely on indirect measures of activity such as hemodynamics resulting in low temporal and spatial resolutions. To improve upon this, optical systems were developed in conjunction with methods to deliver near-IR voltage-sensitive dye (VSD) to provide activity-dependent optical contrast to establish a clinical tool to facilitate direct monitoring of neuron depolarization through the intact skull. Following the previously developed VSD delivery protocol through the blood-brain barrier, IR-780 perchlorate VSD concentrations in the brain were varied and stimulus-evoked responses were observed. In this paper, a range of optimal VSD tissue concentrations was established that maximized fluorescence fractional change for detection of membrane potential responses to external stimuli through a series of phantom, in vitro, ex vivo, and in vivo experiments in mouse models.

scholarly journals Intraoperative ureter identification with a novel fluorescent catheter

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Manuel Barberio ◽  
Mahdi Al-Taher ◽  
Eric Felli ◽  
Anila Hoskere Ashoka ◽  
Jacques Marescaux ◽  
...  
Keyword(s):  
Abdominal Surgery ◽  
Fluorescence Intensity ◽  
Near Infrared ◽  
Fluorescent Imaging ◽  
Human Cadaver ◽  
Fluorescent Material ◽  
Ureteral Injuries ◽  
Normal Light ◽  
Ureteral Peristalsis

AbstractIatrogenic ureteral injuries (IUI) occur in 0.5–1.3% of cases during abdominal surgery. If not recognized intraoperatively, IUI increase morbidity/mortality. A universally accepted method to prevent IUI is lacking. Near-infrared fluorescent imaging (NIRF), penetrating deeper than normal light within the tissue, might be useful, therefore ureter visualization combining NIRF with special dyes (i.e. IRDye 800BK) is promising. Aim of this work is to evaluate the detection of ureters using stents coated with a novel biocompatible fluorescent material (NICE: near-infrared coating of equipment), during laparoscopy. female pigs underwent placement of NICE-coated stents (NS). NIRF was performed, and fluorescence intensity (FI) was computed. Successively, 0.15 mg/kg of IRDye 800BK was administered intravenously, and FI was computed at different timepoints. Ureter visualization using NS only was further assessed in a human cadaver. Both methods allowed in vivo ureter visualization, with equal FI. However, NS were constantly visible whereas IRDye 800BK allowed visualization exclusively during the ureteral peristaltic phases. In the human cadaver, NS provided excellent ureter visualization in its natural anatomical position. NS provided continuous ureteral visualization with similar FI as the IRDye 800BK, which exclusively allowed intermittent visualization, dependent on ureteral peristalsis. NS might prove useful to visualize ureters intraoperatively, potentially preventing IUI.

Biomedical applications: Pitfalls in the practice

1994 ◽  
Vol 52 ◽  
pp. 378-379
Author(s):  
J. D. Shelburne ◽  
Peter Ingram ◽  
Victor L. Roggli ◽  
Ann LeFurgey
Keyword(s):  
Operating Room ◽  
Liquid Nitrogen ◽  
Lung Tissue ◽  
Biomedical Applications ◽  
Microprobe Analysis ◽  
Tissue Sample ◽  
Cellular Level ◽  
Tissue Samples ◽  
Asbestos Fibers

At present most medical microprobe analysis is conducted on insoluble particulates such as asbestos fibers in lung tissue. Cryotechniques are not necessary for this type of specimen. Insoluble particulates can be processed conventionally. Nevertheless, it is important to emphasize that conventional processing is unacceptable for specimens in which electrolyte distributions in tissues are sought. It is necessary to flash-freeze in order to preserve the integrity of electrolyte distributions at the subcellular and cellular level. Ideally, biopsies should be flash-frozen in the operating room rather than being frozen several minutes later in a histology laboratory. Electrolytes will move during such a long delay. While flammable cryogens such as propane obviously cannot be used in an operating room, liquid nitrogen-cooled slam-freezing devices or guns may be permitted, and are the best way to achieve an artifact-free, accurate tissue sample which truly reflects the in vivo state. Unfortunately, the importance of cryofixation is often not understood. Investigators bring tissue samples fixed in glutaraldehyde to a microprobe laboratory with a request for microprobe analysis for electrolytes.

Fluorescent proteins for in vivo imaging, where's the biliverdin?

2020 ◽  
Vol 48 (6) ◽  
pp. 2657-2667
Author(s):  
Felipe Montecinos-Franjola ◽  
John Y. Lin ◽  
Erik A. Rodriguez
Keyword(s):  
Hydrogen Peroxide ◽  
In Vivo Imaging ◽  
Near Infrared ◽  
Fluorescent Protein ◽  
Fluorescent Proteins ◽  
Fluorescent Imaging ◽  
Infrared Light ◽  
Red Fluorescent Protein ◽  
Color Palette

Noninvasive fluorescent imaging requires far-red and near-infrared fluorescent proteins for deeper imaging. Near-infrared light penetrates biological tissue with blood vessels due to low absorbance, scattering, and reflection of light and has a greater signal-to-noise due to less autofluorescence. Far-red and near-infrared fluorescent proteins absorb light &gt;600 nm to expand the color palette for imaging multiple biosensors and noninvasive in vivo imaging. The ideal fluorescent proteins are bright, photobleach minimally, express well in the desired cells, do not oligomerize, and generate or incorporate exogenous fluorophores efficiently. Coral-derived red fluorescent proteins require oxygen for fluorophore formation and release two hydrogen peroxide molecules. New fluorescent proteins based on phytochrome and phycobiliproteins use biliverdin IXα as fluorophores, do not require oxygen for maturation to image anaerobic organisms and tumor core, and do not generate hydrogen peroxide. The small Ultra-Red Fluorescent Protein (smURFP) was evolved from a cyanobacterial phycobiliprotein to covalently attach biliverdin as an exogenous fluorophore. The small Ultra-Red Fluorescent Protein is biophysically as bright as the enhanced green fluorescent protein, is exceptionally photostable, used for biosensor development, and visible in living mice. Novel applications of smURFP include in vitro protein diagnostics with attomolar (10−18 M) sensitivity, encapsulation in viral particles, and fluorescent protein nanoparticles. However, the availability of biliverdin limits the fluorescence of biliverdin-attaching fluorescent proteins; hence, extra biliverdin is needed to enhance brightness. New methods for improved biliverdin bioavailability are necessary to develop improved bright far-red and near-infrared fluorescent proteins for noninvasive imaging in vivo.

Optical imaging probes and their potential contribution to radiotracer development

2016 ◽  
Vol 55 (02) ◽  
pp. 51-62 ◽  
Author(s):  
S. Hermann ◽  
M. Schäfers ◽  
C. Höltke ◽  
A. Faust
Keyword(s):  
Optical Imaging ◽  
Fluorescent Dyes ◽  
Great Influence ◽  
Potential Contribution ◽  
Preclinical Evaluation ◽  
Guided Surgery ◽  
Fluorescence Guided Surgery ◽  
Imaging Probes ◽  
Unspecific Binding

SummaryOptical imaging has long been considered a method for histological or microscopic investigations. Over the last 15 years, however, this method was applied for preclinical molecular imaging and, just recently, was also able to show its principal potential for clinical applications (e.g. fluorescence-guided surgery). Reviewing the development and preclinical evaluation of new fluorescent dyes and target-specific dye conjugates, these often show characteristic patterns of their routes of excretion and biodistribution, which could also be interesting for the development and optimization of radiopharmaceuticals. Especially ionic charges show a great influence on biodistribution and netcharge and charge-distribution on a conjugate often determines unspecific binding or background signals in liver, kidney or intestine, and other organs.Learning from fluorescent probe behaviour in vivo and translating this knowledge to radio-pharmaceuticals might be useful to further optimize emerging and existing radiopharmaceuticals with respect to their biodistribution and thereby availability for binding to their targets.

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