scholarly journals iRFP (near-infrared fluorescent protein) imaging of subcutaneous and deep tissue tumours in mice highlights differences between imaging platforms

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
C. Hall ◽  
Y. von Grabowiecki ◽  
S. P. Pearce ◽  
C. Dive ◽  
S. Bagley ◽  
...  
Keyword(s):  
Cancer Biology ◽  
Near Infrared ◽  
Fluorescent Protein ◽  
Dynamic Range ◽  
Fluorescent Proteins ◽  
Biological Tissues ◽  
The Body ◽  
Nsg Mice ◽  
Infrared Fluorescent Protein

Abstract Background In vivo imaging using fluorescence is used in cancer biology for the detection, measurement and monitoring of tumours. This can be achieved with the expression of fluorescent proteins such as iRFP, which emits light at a wavelength less attenuated in biological tissues compared to light emitted by other fluorescent proteins such as GFP or RFP. Imaging platforms capable of detecting fluorescent tumours in small animals have been developed but studies comparing the performance of these platforms are scarce. Results Through access to three platforms from Xenogen, Bruker and Li-Cor, we compared their ability to detect iRFP-expressing subcutaneous tumours as well as tumours localised deeper within the body of female NSG mice. Each platform was paired with proprietary software for image analyse, but the output depends on subjective decisions from the user. To more objectively compare platforms, we developed an ‘in house’ software-based approach which results in lower measured variability between mice. Conclusions Our comparisons showed that all three platforms allowed for reliable detection and monitoring of subcutaneous iRFP tumour growth. The biggest differences between platforms became apparent when imaging deeper tumours with the Li-Cor platform detecting most tumours and showing the highest dynamic range.

scholarly journals Quantitative assessment of near-infrared fluorescent proteins

2021 ◽  
Author(s):  
Kiryl Piatkevich ◽  
Hanbin Zhang ◽  
Stavrini Papadaki ◽  
Xiaoting Sun ◽  
Luxia Yao ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Quantitative Assessment ◽  
Near Infrared ◽  
Fluorescent Protein ◽  
Fluorescent Proteins ◽  
Oligomeric State ◽  
C Elegans ◽  
The Right ◽  
Infrared Fluorescent Protein

Abstract Recent progress in fluorescent protein development has generated a large diversity of near-infrared fluorescent proteins, which are rapidly becoming popular probes for a variety of imaging applications. To assist end-users with a selection of the right near-infrared fluorescent protein for a given application, we will conduct a quantitative assessment of intracellular brightness, photostability, and oligomeric state of 19 near-infrared fluorescent proteins in cultured mammalian cells. The top-performing proteins will be further validated for in vivo imaging of neurons in C. elegans, zebrafish, and mice. We will also assess the applicability of the selected NIR FPs for expansion microscopy and two-photon imaging.

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 >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.

scholarly journals Fluorescence Phenomena in Amyloid and Amyloidogenic Bionanostructures

Crystals ◽  
2020 ◽  
Vol 10 (8) ◽  
pp. 668 ◽  
Author(s):  
B. Apter ◽  
N. Lapshina ◽  
H. Barhom ◽  
B. Fainberg ◽  
A. Handelman ◽  
...  
Keyword(s):  
Amyloid Fibrils ◽  
Fluorescent Protein ◽  
Fluorescent Proteins ◽  
Protein Structures ◽  
Visible Spectrum ◽  
Biological Tissues ◽  
New Generation ◽  
Β Sheet

Nanoscale optical labeling is an advanced bioimaging tool. It is mostly based on fluorescence (FL) phenomena and enables the visualization of single biocells, bacteria, viruses, and biological tissues, providing monitoring of functional biosystems in vitro and in vivo, and the imaging-guided transportation of drug molecules. There is a variety of FL biolabels such as organic molecular dyes, genetically encoded fluorescent proteins (green fluorescent protein and homologs), semiconductor quantum dots, carbon dots, plasmonic metal gold-based nanostructures and more. In this review, a new generation of FL biolabels based on the recently found biophotonic effects of visible FL are described. This intrinsic FL phenomenon is observed in any peptide/protein materials folded into β-sheet secondary structures, irrespective of their composition, complexity, and origin. The FL effect has been observed both in natural amyloid fibrils, associated with neurodegenerative diseases (Alzheimer’s, Parkinson’s, and more), and diverse synthetic peptide/protein structures subjected to thermally induced biological refolding helix-like→β-sheet. This approach allowed us to develop a new generation of FL peptide/protein bionanodots radiating multicolor, tunable, visible FL, covering the entire visible spectrum in the range of 400–700 nm. Newly developed biocompatible nanoscale biomarkers are considered as a promising tool for emerging precise biomedicine and advanced medical nanotechnologies (high-resolution bioimaging, light diagnostics, therapy, optogenetics, and health monitoring).

scholarly journals Photoacoustic imaging of the near-infrared fluorescent protein iRFP in vivo

2012 ◽  
Author(s):  
Arie Krumholz ◽  
Grigory S. Filonov ◽  
Jun Xia ◽  
Junjie Yao ◽  
Vladislav V. Verkhusha ◽  
...  
Keyword(s):  
Near Infrared ◽  
Fluorescent Protein ◽  
Photoacoustic Imaging ◽  
Infrared Fluorescent Protein

scholarly journals Bright and stable near-infrared fluorescent protein for in vivo imaging

2011 ◽  
Vol 29 (8) ◽  
pp. 757-761 ◽  
Author(s):  
Grigory S Filonov ◽  
Kiryl D Piatkevich ◽  
Li-Min Ting ◽  
Jinghang Zhang ◽  
Kami Kim ◽  
...  
Keyword(s):  
In Vivo Imaging ◽  
Near Infrared ◽  
Fluorescent Protein ◽  
Infrared Fluorescent Protein

scholarly journals Near-Infrared Genetically Encoded Positive Calcium Indicator Based on GAF-FP Bacterial Phytochrome

2019 ◽  
Vol 20 (14) ◽  
pp. 3488 ◽  
Author(s):  
Oksana M. Subach ◽  
Natalia V. Barykina ◽  
Konstantin V. Anokhin ◽  
Kiryl D. Piatkevich ◽  
Fedor V. Subach
Keyword(s):  
Calcium Ions ◽  
Mammalian Cells ◽  
Near Infrared ◽  
Fluorescent Protein ◽  
Dynamic Range ◽  
Cultured Cells ◽  
Infrared Region ◽  
Calcium Indicator

A variety of genetically encoded calcium indicators are currently available for visualization of calcium dynamics in cultured cells and in vivo. Only one of them, called NIR-GECO1, exhibits fluorescence in the near-infrared region of the spectrum. NIR-GECO1 is engineered based on the near-infrared fluorescent protein mIFP derived from bacterial phytochromes. However, NIR-GECO1 has an inverted response to calcium ions and its excitation spectrum is not optimal for the commonly used 640 nm lasers. Using small near-infrared bacterial phytochrome GAF-FP and calmodulin/M13-peptide pair, we developed a near-infrared calcium indicator called GAF-CaMP2. In vitro, GAF-CaMP2 showed a positive response of 78% and high affinity (Kd of 466 nM) to the calcium ions. It had excitation and emission maxima at 642 and 674 nm, respectively. GAF-CaMP2 had a 2.0-fold lower brightness, 5.5-fold faster maturation and lower pH stability compared to GAF-FP in vitro. GAF-CaMP2 showed 2.9-fold higher photostability than smURFP protein. The GAF-CaMP2 fusion with sfGFP demonstrated a ratiometric response with a dynamic range of 169% when expressed in the cytosol of mammalian cells in culture. Finally, we successfully applied the ratiometric version of GAF-CaMP2 for the simultaneous visualization of calcium transients in three organelles of mammalian cells using four-color fluorescence microscopy.

scholarly journals Non-invasive in vivo imaging of UCP1 expression in live mice via near-infrared fluorescent protein iRFP720

PLoS ONE ◽  
2019 ◽  
Vol 14 (11) ◽  
pp. e0225213 ◽  
Author(s):  
Aya Fukuda ◽  
Shiho Honda ◽  
Norie Fujioka ◽  
Yuya Sekiguchi ◽  
Seiya Mizuno ◽  
...  
Keyword(s):  
In Vivo Imaging ◽  
Near Infrared ◽  
Fluorescent Protein ◽  
Non Invasive ◽  
Ucp1 Expression ◽  
Infrared Fluorescent Protein

scholarly journals Synthesis and Characterization of GdVO4:Nd Near-Infrared Phosphors for Optical Time-Gated In Vivo Imaging

Materials ◽  
2020 ◽  
Vol 13 (16) ◽  
pp. 3564
Author(s):  
Ben Nimmegeers ◽  
Ewoud Cosaert ◽  
Tecla Carbonati ◽  
Daniela Meroni ◽  
Dirk Poelman
Keyword(s):  
Decay Time ◽  
Near Infrared ◽  
Imaging Techniques ◽  
Medical Personnel ◽  
Biological Tissues ◽  
The Body ◽  
Transparency Window ◽  
Optical Time ◽  
Gated Imaging

Many medical imaging techniques use some form of ionizing radiation. This radiation is not only potentially harmful for the patient, but also for the medical personnel. An alternative imaging technique uses near-infrared (NIR) emitting luminescent particles as tracers. If the luminescent probes are excited inside the body, autofluorescence from the biological tissues is also induced. This problem can be circumvented by using time-gated imaging. Hereby, the light collection only starts when the fluorescence of the tissue has decayed. This requires particles showing both excitation and emission in the near-infrared and a long decay time so that they can be used in time-gated imaging. In this work, Nd-doped GdVO4 NIR emitting particles were prepared using solid state reaction. Particles could be efficiently excited at 808 nm, right in the first transparency window for biological tissues, emitted in the second transparency window at around 1064 nm, and showed a decay time of the order of 70 μs, sufficiently long for time-gating. By using a Gd-containing host, these particles could be ideally suited for multimodal optical/magnetic imaging after size reduction and surface functionalization.

scholarly journals GAF-CaMP3–sfGFP, An Enhanced Version of the Near-Infrared Genetically Encoded Positive Phytochrome-Based Calcium Indicator for the Visualization of Neuronal Activity

2020 ◽  
Vol 21 (18) ◽  
pp. 6883
Author(s):  
Oksana M. Subach ◽  
Fedor V. Subach
Keyword(s):  
Calcium Ions ◽  
Near Infrared ◽  
Fluorescent Protein ◽  
Dynamic Range ◽  
Fluorescent Proteins ◽  
Spectral Characteristics ◽  
Confocal Imaging ◽  
Neuronal Cultures ◽  
Decay Kinetics ◽  
Calcium Indicator

The first generation of near-infrared, genetically encoded calcium indicators (NIR-GECIs) was developed from bacterial phytochrome-based fluorescent proteins that utilize biliverdin (BV) as the chromophore moiety. However, NIR-GECIs have some main drawbacks such as either an inverted response to calcium ions (in the case of NIR-GECO1) or a limited dynamic range and a lack of data about their application in neurons (in the case of GAF-CaMP2–superfolder green fluorescent protein (sfGFP)). Here, we developed an enhanced version of the GAF-CaMP2–sfGFP indicator, named GAF-CaMP3–sfGFP. The GAF-CaMP3–sfGFP demonstrated spectral characteristics, molecular brightness, and a calcium affinity similar to the respective characteristics for its progenitor, but a 2.9-fold larger DF/F response to calcium ions. As compared to GAF-CaMP2–sfGFP, in cultured HeLa cells, GAF-CaMP3–sfGFP had similar brightness but a 1.9-fold larger DF/F response to the elevation of calcium ions levels. Finally, we successfully utilized the GAF-CaMP3–sfGFP for the monitoring of the spontaneous and stimulated activity of neuronal cultures and compared its performance with the R-GECO1 indicator using two-color confocal imaging. In the cultured neurons, GAF-CaMP3–sfGFP showed a linear DF/F response in the range of 0–20 APs and in this range demonstrated a 1.4-fold larger DF/F response but a 1.3- and 2.4-fold slower rise and decay kinetics, respectively, as compared to the same parameters for the R-GECO1 indicator.

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