Red Fluorescent Protein

14,000,000 Leading Edge Experts on the ideXlab platform

Scan Science and Technology

Contact Leading Edge Experts & Companies

Scan Science and Technology

Contact Leading Edge Experts & Companies

The Experts below are selected from a list of 360 Experts worldwide ranked by ideXlab platform

Robert M. Hoffman - One of the best experts on this subject based on the ideXlab platform.

  • analysis of stroma labeling during multiple passage of a sarcoma imageable patient derived orthotopic xenograft ipdox in Red Fluorescent Protein transgenic nude mice
    Journal of Cellular Biochemistry, 2017
    Co-Authors: Tasuku Kiyuna, Yasunori Tome, Kentaro Miyake, Kei Kawaguchi, Takashi Murakami, Kentaro Igarashi, Fuminori Kanaya, Arun S Singh, Fritz C Eilber, Robert M. Hoffman
    Abstract:

    A patient-derived orthotopic xenograft (PDOX) model of undifferentiated pleomorphic sarcoma (UPS) was previously established that acquiRed Red Fluorescent Protein (RFP)-expressing stroma by growth in an RFP transgenic nude mouse. In the present study, an imageable PDOX model (iPDOX) of UPS was established by orthotopic implantation in the biceps femoris of transgenic RFP nude mice. After the tumors grew to a diameter of 10 mm, they were harvested and the brightest portion of the tumors were subsequently orthotopically transplanted to both RFP and non-coloRed nude mice. The UPS PDOX tumor was again transplanted to RFP transgenic and non-coloRed nude mice, and finally a 3rd passage was made in the same manner. Five UPS tumors from each passage in both RFP and non-coloRed mouse models were harvested. The FV1,000 confocal microscope was used to visualize and quantitate the RFP area of the resected tumors. The average percent Fluorescent area in the first passage of RFP mice was 34 ± 22%; in the second passage, 34 ± 20%; and 36 ± 11% in the third passage of RFP transgenic nude mice. The average tumor RFP area in the first passage from RFP mice to non-coloRed mice was 20 ± 7%; in the second passage, 28 ± 11%; in the third passage was 27 ± 13%. The present results demonstrate the extensive and stable acquisition of stroma by the UPS-tumor growing orthotopically in transgenic RFP nude mice (iPDOX). This model can be used for screening for effective drugs for individual patients and drug discovery. J. Cell. Biochem. 118: 3367-3371, 2017. © 2017 Wiley Periodicals, Inc.

  • abstract 4221 imaging formation of the tumor microenvironment in orthotopic liver tumors in Red Fluorescent Protein rfp transgenic nude mice
    Cancer Research, 2016
    Co-Authors: Atsushi Suetsugu, Michael Bouvet, Yukihiko Hiroshima, Takuro Matsumoto, Kosuke Hasegawa, Miki Nakamura, Masahito Shimizu, Shigetoyo Saji, Hisataka Moriwaki, Robert M. Hoffman
    Abstract:

    We report here dual-color imaging of the formation of the TME in orthotopic liver cancer in transgenic Red Fluorescent Protein (RFP) nude mice, which express RFP in all organs. Non-coloRed HUH-7 human hepatoma cells were injected in the spleen of RFP nude mice to establish an orthotopic liver cancer model. TME formation in the liver tumor was observed using the Olympus OV100 small animal fluorescence imaging system. Non-coloRed liver cancer cells formed colonies in the liver 28 days after cell transplantation to the spleen. In the liver, RFP host-mouse cells accumulated around and in the liver tumor as visualized by fluorescence imaging. A desmin and sirus-Red positive area increased around and within the liver tumor over time. These results indicate cancer-associated fibroblasts (CAFs) were recruited by the liver metastasis suggesting that CAFs, along with the angiogenic tumor blood vessels, could serve as visible therapeutic targets. Citation Format: Atsushi Suetsugu, Yukihiko Hiroshima, Takuro Matsumoto, Kosuke Hasegawa, Miki Nakamura, Masahito Shimizu, Shigetoyo Saji, Hisataka Moriwaki, Michael Bouvet, Robert M. Hoffman. Imaging formation of the tumor microenvironment in orthotopic liver tumors in Red Fluorescent Protein (RFP) transgenic nude mice. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4221.

  • combination efficacy of astragalus membranaceus and curcuma wenyujin at different stages of tumor progression in an imageable orthotopic nude mouse model of metastatic human ovarian cancer expressing Red Fluorescent Protein
    Anticancer Research, 2015
    Co-Authors: Gang Yin, Robert M. Hoffman, Decai Tang, Jianguo Dai, Min Liu, Y U Sun, Zhijian Yang, Shuo Zhang, Xiuxia Guo
    Abstract:

    Background/aim The present study determined the efficacy of extracts of Astragalus membranaceus (AM) and Curcuma wenyujin (CW), a traditional Chinese medicine herbal mixture, at different tumor stages of an orthotopic nude mouse model of human ovarian cancer expressing Red Fluorescent Protein. Materials and methods The tumor-bearing mice were treated with cisplatinum (CDDP), AM, CW, or a combination of AM and CW in each of three tumor stages, using the same regimen. Group 1 received saline as negative control. Group 2 received CDDP i.p. as positive control with a dose of 2 mg/kg, every three days. Group 3 received AM daily via oral gavage, at a dose of 9120 mg/kg. Group 4 received CW daily via oral gavage, at a dose of 4560 mg/kg. Groups 5, 6 and 7 received combinations of AM and CW daily via oral gavage at low (AM, 2280 mg/kg; CW, 1140 mg/kg), medium (AM, 4560 mg/kg; CW 2280 mg/kg), and high (AM, 9120 mg/kg; CW, 4560 mg/kg) doses. The expression of angiogenesis- and apoptosis-related genes in the tumors were analyzed by immunohistochemistry for matrix metalloProteinase 2 (MMP-2), vascular endothelial growth factor (VEGF) fibroblast growth factor 2 (FGF-2), B-cell lymphoma 2 (Bcl-2) and cyclooxygenase 2 (Cox-2), and by polymerase chain reaction for MMP-2, FGF-2 and Bcl-2. Results CDDP, AM, and its combination with CW-induced significant growth inhibition of Stage I tumors. Strong efficacy of the combination of AM and CW at high dose was observed. Monotherapy with CDDP, AM, CW, and the combination treatments did not significantly inhibit Stage II and III tumors. The expression of MMP-2, VEGF, FGF-2, and Cox-2 was significantly Reduced in Stage I tumors treated with AM, CW, and their combination, suggesting a possible role of these angiogenesis- and apoptosis-related genes in the observed efficacy of the agents tested. Conclusion This study is the first report on the efficacy of anticancer agents at different stages of ovarian cancer in an orthotopic mouse model. As the tumor progressed, it became treatment-resistant, similar to the clinical situation, further demonstrating the utility of the model and the need for agents acrtive in advanced-stage ovarian cancer.

  • early reporting of apoptosis by real time imaging of cancer cells labeled with green Fluorescent Protein in the nucleus and Red Fluorescent Protein in the cytoplasm
    Anticancer Research, 2015
    Co-Authors: Meng Yang, Ping Jiang, Robert M. Hoffman
    Abstract:

    BACKGROUND/AIM We previously developed PC-3 human prostate cancer cells expressing Red Fluorescent Protein (RFP) in the cytoplasm and green Fluorescent Protein (GFP) linked to histone H2B expressed in the nucleus. We demonstrate in the present report the use of these dual-color cells for early detection of apoptosis in the presence of cancer chemotherapy agents. MATERIALS AND METHODS Induction of apoptosis was observed by real-time imaging of cytoplasmic and nuclear size and shape changes and nuclear fragmentation using fluorescence microscopy. Apoptosis was also detected by measuring DNA fragmentation. The cancer chemotherapy agents paclitaxel and vinblastine were used for induction of apoptosis. RESULTS When the PC-3 dual-color cells were treated with paclitaxel or vinblastine, cytoplasmic and nuclear size and shape changes and nuclear fragmentation were observed by 24 hours. The paclitaxel-treated PC-3 dual-color cells exhibited ring-like structures formed by the fragmented nuclei, which could be brightly visualized by H2B-GFP fluorescence. Apoptosis was also detected by the dual-color PC-3 cells by 24 hours when treated with vinblastine. However, no nuclear ring-like structures were formed in the PC-3 cells by vinblastine treatment. In contrast, DNA fragmentation could not be observed in PC-3 cells until 48 hours after exposure to paclitaxel. CONCLUSION Dual-color PC-3 cells can serve as a simple real-time early reporter of apoptosis and as a screen for novel cancer therapeutics or genotoxic agents. The dual-color cell real-time imaging assay is a more sensitive and earlier reporter for apoptosis than the DNA fragmentation assay.

  • Selective Antimetastatic Activity of Cytosine Analog CS-682 in a Red Fluorescent Protein Orthotopic Model of Pancreatic Cancer 1
    2013
    Co-Authors: Matthew H. Katz, Michael Bouvet, Shinako Takimoto, Daniel Spivack, Abdool R. Moossa, Robert M. Hoffman
    Abstract:

    In this study we demonstrate the ability of a novel, p.o.-administeRed cytosine analogue, CS-682, to effectively prolong survival and inhibit metastatic growth in an imageable orthotopic mouse model of pancreatic cancer. MIA-PaCa-2-RFP pancreatic cancer cells were transduced with the Discosoma Red Fluorescent Protein (RFP) and orthotopically implanted onto the pancreas of nude mice. Tumor RFP fluorescence facilitated real-time, sequential imaging, and quantification of primary and metastatic growth and dissemination in vivo. Mice were treated with various p.o. doses of CS-682 on a five times per week schedule until death. At a dose of 40 mg/kg, CS-682 prolonged survival compaRed with untreated animals (median survival 35 days versus 17 days; P � 0.0008). At nontoxic doses, CS-682 effectively suppressed the rate of primary tumor growth. CS-682 also decreased the development of malignant ascites and the formation of metastases, which were Reduced significantly in number i

Roger Y Tsien - One of the best experts on this subject based on the ideXlab platform.

  • a far Red Fluorescent Protein evolved from a cyanobacterial phycobiliProtein
    Nature Methods, 2016
    Co-Authors: Erik A Rodriguez, Roger Y Tsien, Geraldine N Tran, Larry A Gross, Jessica L Crisp, Xiaokun Shu, John Y Lin
    Abstract:

    A bright and photostable far-Red Fluorescent Protein, smURFP, was developed from a cyanobacterial phycobiliProtein. smURFP uniquely binds a highly cell-permeable biliverdin derivative to obtain fluorescence brightness comparable to that of eGFP in cells.

  • functional and structural characterization of a new monomeric far Red Fluorescent Protein
    Biophysical Journal, 2010
    Co-Authors: Michael Z Lin, Michael R Mckeown, Tom Alber, Roger Y Tsien
    Abstract:

    Fluorescent Proteins have become valuable tools for biomedical research as Protein tags, reporters of gene expression, biosensor components, and cell lineage tracers. However, applications of Fluorescent Proteins for deep tissue imaging have been constrained by the opacity of tissues to excitation light below 600 nm, due to absorbance by hemoglobin. Fluorescent Proteins that excite efficiently in the “optical window” above 600 nm are therefore highly desirable. We report here the evolution of a far-Red Fluorescent Protein with peak excitation at 600 nm and peak emission at 650 nm. This, Neptune, performs well in imaging deep tissues in living mice. The crystal structure of Neptune reveals novel mechanisms for Red-shifting, including the acquisition of a new hydrogen bond with the chromophore. Neptune may serve as the basis for Fluorescent indicators or FRET reporters that are more compatible with deep tissue imaging.

  • improved monomeric Red orange and yellow Fluorescent Proteins derived from discosoma sp Red Fluorescent Protein
    Nature Biotechnology, 2004
    Co-Authors: Nathan C. Shaner, Robert E. Campbell, Paul Steinbach, Ben N G Giepmans, Amy E Palmer, Roger Y Tsien
    Abstract:

    Improved monomeric Red, orange and yellow Fluorescent Proteins derived from Discosoma sp. Red Fluorescent Protein

  • a monomeric Red Fluorescent Protein
    Proceedings of the National Academy of Sciences of the United States of America, 2002
    Co-Authors: Robert E. Campbell, Paul Steinbach, Amy E Palmer, Oded Tour, Geoffrey S Baird, David A Zacharias, Roger Y Tsien
    Abstract:

    All coelenterate Fluorescent Proteins cloned to date display some form of quaternary structure, including the weak tendency of Aequorea green Fluorescent Protein (GFP) to dimerize, the obligate dimerization of Renilla GFP, and the obligate tetramerization of the Red Fluorescent Protein from Discosoma (DsRed). Although the weak dimerization of Aequorea GFP has not impeded its acceptance as an indispensable tool of cell biology, the obligate tetramerization of DsRed has greatly hindeRed its use as a genetically encoded fusion tag. We present here the stepwise evolution of DsRed to a dimer and then either to a genetic fusion of two copies of the Protein, i.e., a tandem dimer, or to a true monomer designated mRFP1 (monomeric Red Fluorescent Protein). Each subunit interface was disrupted by insertion of arginines, which initially crippled the resulting Protein, but Red fluorescence could be rescued by random and directed mutagenesis totaling 17 substitutions in the dimer and 33 in mRFP1. Fusions of the gap junction Protein connexin43 to mRFP1 formed fully functional junctions, whereas analogous fusions to the tetramer and dimer failed. Although mRFP1 has somewhat lower extinction coefficient, quantum yield, and photostability than DsRed, mRFP1 matures >10 times faster, so that it shows similar brightness in living cells. In addition, the excitation and emission peaks of mRFP1, 584 and 607 nm, are ≈25 nm Red-shifted from DsRed, which should confer greater tissue penetration and spectral separation from autofluorescence and other Fluorescent Proteins.

  • the structure of the chromophore within dsRed a Red Fluorescent Protein from coral
    Proceedings of the National Academy of Sciences of the United States of America, 2000
    Co-Authors: Larry A Gross, Geoffrey S Baird, Ross C Hoffman, Kim K Baldridge, Roger Y Tsien
    Abstract:

    Abstract DsRed, a brilliantly Red Fluorescent Protein, was recently cloned from Discosoma coral by homology to the green Fluorescent Protein (GFP) from the jellyfish Aequorea. A core question in the biochemistry of DsRed is the mechanism by which the GFP-like 475-nm excitation and 500-nm emission maxima of immature DsRed are Red-shifted to the 558-nm excitation and 583-nm emission maxima of mature DsRed. After digestion of mature DsRed with lysyl endopeptidase, high-resolution mass spectra of the purified chromophore-bearing peptide reveal that some of the molecules have lost 2 Da relative to the peptide analogously prepaRed from a mutant, K83R, that stays green. Tandem mass spectrometry indicates that the bond between the alpha-carbon and nitrogen of Gln-66 has been dehydrogenated in DsRed, extending the GFP chromophore by forming —C⩵N—C⩵O at the 2-position of the imidazolidinone. This acylimine substituent quantitatively accounts for the Red shift according to quantum mechanical calculations. Reversible hydration of the C⩵N bond in the acylimine would explain why denaturation shifts mature DsRed back to a GFP-like absorbance. The C⩵N bond hydrolyses upon boiling, explaining why DsRed shows two fragment bands on SDS/PAGE. This assay suggests that conversion from green to Red chromophores remains incomplete even after prolonged aging.

Ahmed S Abdelfattah - One of the best experts on this subject based on the ideXlab platform.

  • a genetically encoded ca2 indicator based on circularly permutated sea anemone Red Fluorescent Protein eqfp578
    BMC Biology, 2018
    Co-Authors: Yi Shen, Ahmed S Abdelfattah, Hod Dana, Ronak Patel, Jamien Shea, Rosana S Molina, Bijal Rawal, Vladimir Rancic
    Abstract:

    Genetically encoded calcium ion (Ca2+) indicators (GECIs) are indispensable tools for measuring Ca2+ dynamics and neuronal activities in vitro and in vivo. Red Fluorescent Protein (RFP)-based GECIs have inherent advantages relative to green Fluorescent Protein-based GECIs due to the longer wavelength light used for excitation. Longer wavelength light is associated with decreased phototoxicity and deeper penetration through tissue. Red GECI can also enable multicolor visualization with blue- or cyan-excitable fluorophores. Here we report the development, structure, and validation of a new RFP-based GECI, K-GECO1, based on a circularly permutated RFP derived from the sea anemone Entacmaea quadricolor. We have characterized the performance of K-GECO1 in cultuRed HeLa cells, dissociated neurons, stem-cell-derived cardiomyocytes, organotypic brain slices, zebrafish spinal cord in vivo, and mouse brain in vivo. K-GECO1 is the archetype of a new lineage of GECIs based on the RFP eqFP578 scaffold. It offers high sensitivity and fast kinetics, similar or better than those of current state-of-the-art indicators, with diminished lysosomal accumulation and minimal blue-light photoactivation. Further refinements of the K-GECO1 lineage could lead to further improved variants with overall performance that exceeds that of the most highly optimized Red GECIs.

  • a genetically encoded ca 2 indicator based on circularly permutated sea anemone Red Fluorescent Protein eqfp578
    BMC Biology, 2018
    Co-Authors: Yi Shen, Ahmed S Abdelfattah, Hod Dana, Ronak Patel, Jamien Shea, Rosana S Molina, Bijal Rawal, Vladimir Rancic
    Abstract:

    Genetically encoded calcium ion (Ca2+) indicators (GECIs) are indispensable tools for measuring Ca2+ dynamics and neuronal activities in vitro and in vivo. Red Fluorescent Protein (RFP)-based GECIs have inherent advantages relative to green Fluorescent Protein-based GECIs due to the longer wavelength light used for excitation. Longer wavelength light is associated with decreased phototoxicity and deeper penetration through tissue. Red GECI can also enable multicolor visualization with blue- or cyan-excitable fluorophores. Here we report the development, structure, and validation of a new RFP-based GECI, K-GECO1, based on a circularly permutated RFP derived from the sea anemone Entacmaea quadricolor. We have characterized the performance of K-GECO1 in cultuRed HeLa cells, dissociated neurons, stem-cell-derived cardiomyocytes, organotypic brain slices, zebrafish spinal cord in vivo, and mouse brain in vivo. K-GECO1 is the archetype of a new lineage of GECIs based on the RFP eqFP578 scaffold. It offers high sensitivity and fast kinetics, similar or better than those of current state-of-the-art indicators, with diminished lysosomal accumulation and minimal blue-light photoactivation. Further refinements of the K-GECO1 lineage could lead to further improved variants with overall performance that exceeds that of the most highly optimized Red GECIs.

  • A genetically encoded Ca2+ indicator based on circularly permutated sea anemone Red Fluorescent Protein eqFP578
    BMC, 2018
    Co-Authors: Yi Shen, Ahmed S Abdelfattah, Hod Dana, Ronak Patel, Jamien Shea, Rosana S Molina, Bijal Rawal, Vladimir Rancic, Yufen Chang
    Abstract:

    Abstract Background Genetically encoded calcium ion (Ca2+) indicators (GECIs) are indispensable tools for measuring Ca2+ dynamics and neuronal activities in vitro and in vivo. Red Fluorescent Protein (RFP)-based GECIs have inherent advantages relative to green Fluorescent Protein-based GECIs due to the longer wavelength light used for excitation. Longer wavelength light is associated with decreased phototoxicity and deeper penetration through tissue. Red GECI can also enable multicolor visualization with blue- or cyan-excitable fluorophores. Results Here we report the development, structure, and validation of a new RFP-based GECI, K-GECO1, based on a circularly permutated RFP derived from the sea anemone Entacmaea quadricolor. We have characterized the performance of K-GECO1 in cultuRed HeLa cells, dissociated neurons, stem-cell-derived cardiomyocytes, organotypic brain slices, zebrafish spinal cord in vivo, and mouse brain in vivo. Conclusion K-GECO1 is the archetype of a new lineage of GECIs based on the RFP eqFP578 scaffold. It offers high sensitivity and fast kinetics, similar or better than those of current state-of-the-art indicators, with diminished lysosomal accumulation and minimal blue-light photoactivation. Further refinements of the K-GECO1 lineage could lead to further improved variants with overall performance that exceeds that of the most highly optimized Red GECIs

  • a genetically encoded ca2 indicator based on circularly permutated sea anemone Red Fluorescent Protein
    bioRxiv, 2017
    Co-Authors: Yi Shen, Ahmed S Abdelfattah, Hod Dana, Ronak Patel, Jamien Shea, Rosana S Molina, Bijal Rawal, Vladimir Rancic, Yufen Chang, Yingche Chen
    Abstract:

    Genetically-encoded calcium ion (Ca2+) indicators (GECIs) are indispensable tools for measuring Ca2+ dynamics and neuronal activities in vitro and in vivo. Red Fluorescent Protein (RFP)-based GECIs enable multicolor visualization with blue or cyan-excitable fluorophores and combined use with blue or cyan-excitable optogenetic actuators. Here we report the development, structure, and validation of a new Red Fluorescent Ca2+ indicator, K-GECO1, based on a circularly permutated RFP derived from the sea anemone Entacmaea quadricolor. We characterized the performance of K-GECO1 in cultuRed HeLa cells, dissociated neurons, stem cell derived cardiomyocytes, organotypic brain slices, zebrafish spinal cord in vivo, and mouse brain in vivo.

  • a fluorogenic Red Fluorescent Protein heterodimer
    Chemistry & Biology, 2012
    Co-Authors: Spencer C Alford, Ahmed S Abdelfattah, Yidan Ding, Robert E. Campbell
    Abstract:

    Summary The expanding repertoire of genetically encoded biosensors constructed from variants of Aequorea victoria green Fluorescent Protein (GFP) enable the imaging of a variety of intracellular biochemical processes. To facilitate the imaging of multiple biosensors in a single cell, we undertook the development of a dimerization-dependent Red Fluorescent Protein (ddRFP) that provides an alternative strategy for biosensor construction. An extensive process of rational engineering and directed Protein evolution led to the discovery of a ddRFP with a K d of 33 μM and a 10-fold increase in fluorescence upon heterodimer formation. We demonstrate that the dimerization-dependent fluorescence of ddRFP can be used for detection of a Protein-Protein interaction in vitro, imaging of the reversible Ca 2+ -dependent association of calmodulin and M13 in live cells, and imaging of caspase-3 activity during apoptosis.

Yi Shen - One of the best experts on this subject based on the ideXlab platform.

  • a genetically encoded ca2 indicator based on circularly permutated sea anemone Red Fluorescent Protein eqfp578
    BMC Biology, 2018
    Co-Authors: Yi Shen, Ahmed S Abdelfattah, Hod Dana, Ronak Patel, Jamien Shea, Rosana S Molina, Bijal Rawal, Vladimir Rancic
    Abstract:

    Genetically encoded calcium ion (Ca2+) indicators (GECIs) are indispensable tools for measuring Ca2+ dynamics and neuronal activities in vitro and in vivo. Red Fluorescent Protein (RFP)-based GECIs have inherent advantages relative to green Fluorescent Protein-based GECIs due to the longer wavelength light used for excitation. Longer wavelength light is associated with decreased phototoxicity and deeper penetration through tissue. Red GECI can also enable multicolor visualization with blue- or cyan-excitable fluorophores. Here we report the development, structure, and validation of a new RFP-based GECI, K-GECO1, based on a circularly permutated RFP derived from the sea anemone Entacmaea quadricolor. We have characterized the performance of K-GECO1 in cultuRed HeLa cells, dissociated neurons, stem-cell-derived cardiomyocytes, organotypic brain slices, zebrafish spinal cord in vivo, and mouse brain in vivo. K-GECO1 is the archetype of a new lineage of GECIs based on the RFP eqFP578 scaffold. It offers high sensitivity and fast kinetics, similar or better than those of current state-of-the-art indicators, with diminished lysosomal accumulation and minimal blue-light photoactivation. Further refinements of the K-GECO1 lineage could lead to further improved variants with overall performance that exceeds that of the most highly optimized Red GECIs.

  • a genetically encoded ca 2 indicator based on circularly permutated sea anemone Red Fluorescent Protein eqfp578
    BMC Biology, 2018
    Co-Authors: Yi Shen, Ahmed S Abdelfattah, Hod Dana, Ronak Patel, Jamien Shea, Rosana S Molina, Bijal Rawal, Vladimir Rancic
    Abstract:

    Genetically encoded calcium ion (Ca2+) indicators (GECIs) are indispensable tools for measuring Ca2+ dynamics and neuronal activities in vitro and in vivo. Red Fluorescent Protein (RFP)-based GECIs have inherent advantages relative to green Fluorescent Protein-based GECIs due to the longer wavelength light used for excitation. Longer wavelength light is associated with decreased phototoxicity and deeper penetration through tissue. Red GECI can also enable multicolor visualization with blue- or cyan-excitable fluorophores. Here we report the development, structure, and validation of a new RFP-based GECI, K-GECO1, based on a circularly permutated RFP derived from the sea anemone Entacmaea quadricolor. We have characterized the performance of K-GECO1 in cultuRed HeLa cells, dissociated neurons, stem-cell-derived cardiomyocytes, organotypic brain slices, zebrafish spinal cord in vivo, and mouse brain in vivo. K-GECO1 is the archetype of a new lineage of GECIs based on the RFP eqFP578 scaffold. It offers high sensitivity and fast kinetics, similar or better than those of current state-of-the-art indicators, with diminished lysosomal accumulation and minimal blue-light photoactivation. Further refinements of the K-GECO1 lineage could lead to further improved variants with overall performance that exceeds that of the most highly optimized Red GECIs.

  • A genetically encoded Ca2+ indicator based on circularly permutated sea anemone Red Fluorescent Protein eqFP578
    BMC, 2018
    Co-Authors: Yi Shen, Ahmed S Abdelfattah, Hod Dana, Ronak Patel, Jamien Shea, Rosana S Molina, Bijal Rawal, Vladimir Rancic, Yufen Chang
    Abstract:

    Abstract Background Genetically encoded calcium ion (Ca2+) indicators (GECIs) are indispensable tools for measuring Ca2+ dynamics and neuronal activities in vitro and in vivo. Red Fluorescent Protein (RFP)-based GECIs have inherent advantages relative to green Fluorescent Protein-based GECIs due to the longer wavelength light used for excitation. Longer wavelength light is associated with decreased phototoxicity and deeper penetration through tissue. Red GECI can also enable multicolor visualization with blue- or cyan-excitable fluorophores. Results Here we report the development, structure, and validation of a new RFP-based GECI, K-GECO1, based on a circularly permutated RFP derived from the sea anemone Entacmaea quadricolor. We have characterized the performance of K-GECO1 in cultuRed HeLa cells, dissociated neurons, stem-cell-derived cardiomyocytes, organotypic brain slices, zebrafish spinal cord in vivo, and mouse brain in vivo. Conclusion K-GECO1 is the archetype of a new lineage of GECIs based on the RFP eqFP578 scaffold. It offers high sensitivity and fast kinetics, similar or better than those of current state-of-the-art indicators, with diminished lysosomal accumulation and minimal blue-light photoactivation. Further refinements of the K-GECO1 lineage could lead to further improved variants with overall performance that exceeds that of the most highly optimized Red GECIs

  • a genetically encoded ca2 indicator based on circularly permutated sea anemone Red Fluorescent Protein
    bioRxiv, 2017
    Co-Authors: Yi Shen, Ahmed S Abdelfattah, Hod Dana, Ronak Patel, Jamien Shea, Rosana S Molina, Bijal Rawal, Vladimir Rancic, Yufen Chang, Yingche Chen
    Abstract:

    Genetically-encoded calcium ion (Ca2+) indicators (GECIs) are indispensable tools for measuring Ca2+ dynamics and neuronal activities in vitro and in vivo. Red Fluorescent Protein (RFP)-based GECIs enable multicolor visualization with blue or cyan-excitable fluorophores and combined use with blue or cyan-excitable optogenetic actuators. Here we report the development, structure, and validation of a new Red Fluorescent Ca2+ indicator, K-GECO1, based on a circularly permutated RFP derived from the sea anemone Entacmaea quadricolor. We characterized the performance of K-GECO1 in cultuRed HeLa cells, dissociated neurons, stem cell derived cardiomyocytes, organotypic brain slices, zebrafish spinal cord in vivo, and mouse brain in vivo.

  • phuji a ph sensitive Red Fluorescent Protein for imaging of exo and endocytosis
    Journal of Cell Biology, 2014
    Co-Authors: Yi Shen, Robert E. Campbell, Morgane Rosendale, David Perrais
    Abstract:

    Fluorescent Proteins with pH-sensitive fluorescence are valuable tools for the imaging of exocytosis and endocytosis. The Aequorea green Fluorescent Protein mutant superecliptic pHluorin (SEP) is particularly well suited to these applications. Here we describe pHuji, a Red Fluorescent Protein with a pH sensitivity that approaches that of SEP, making it amenable for detection of single exocytosis and endocytosis events. To demonstrate the utility of the pHuji plus SEP pair, we perform simultaneous two-color imaging of clathrin-mediated internalization of both the transferrin receptor and the β2 adrenergic receptor. These experiments reveal that the two receptors are differentially sorted at the time of endocytic vesicle formation.

Vladimir Rancic - One of the best experts on this subject based on the ideXlab platform.

  • a genetically encoded ca2 indicator based on circularly permutated sea anemone Red Fluorescent Protein eqfp578
    BMC Biology, 2018
    Co-Authors: Yi Shen, Ahmed S Abdelfattah, Hod Dana, Ronak Patel, Jamien Shea, Rosana S Molina, Bijal Rawal, Vladimir Rancic
    Abstract:

    Genetically encoded calcium ion (Ca2+) indicators (GECIs) are indispensable tools for measuring Ca2+ dynamics and neuronal activities in vitro and in vivo. Red Fluorescent Protein (RFP)-based GECIs have inherent advantages relative to green Fluorescent Protein-based GECIs due to the longer wavelength light used for excitation. Longer wavelength light is associated with decreased phototoxicity and deeper penetration through tissue. Red GECI can also enable multicolor visualization with blue- or cyan-excitable fluorophores. Here we report the development, structure, and validation of a new RFP-based GECI, K-GECO1, based on a circularly permutated RFP derived from the sea anemone Entacmaea quadricolor. We have characterized the performance of K-GECO1 in cultuRed HeLa cells, dissociated neurons, stem-cell-derived cardiomyocytes, organotypic brain slices, zebrafish spinal cord in vivo, and mouse brain in vivo. K-GECO1 is the archetype of a new lineage of GECIs based on the RFP eqFP578 scaffold. It offers high sensitivity and fast kinetics, similar or better than those of current state-of-the-art indicators, with diminished lysosomal accumulation and minimal blue-light photoactivation. Further refinements of the K-GECO1 lineage could lead to further improved variants with overall performance that exceeds that of the most highly optimized Red GECIs.

  • a genetically encoded ca 2 indicator based on circularly permutated sea anemone Red Fluorescent Protein eqfp578
    BMC Biology, 2018
    Co-Authors: Yi Shen, Ahmed S Abdelfattah, Hod Dana, Ronak Patel, Jamien Shea, Rosana S Molina, Bijal Rawal, Vladimir Rancic
    Abstract:

    Genetically encoded calcium ion (Ca2+) indicators (GECIs) are indispensable tools for measuring Ca2+ dynamics and neuronal activities in vitro and in vivo. Red Fluorescent Protein (RFP)-based GECIs have inherent advantages relative to green Fluorescent Protein-based GECIs due to the longer wavelength light used for excitation. Longer wavelength light is associated with decreased phototoxicity and deeper penetration through tissue. Red GECI can also enable multicolor visualization with blue- or cyan-excitable fluorophores. Here we report the development, structure, and validation of a new RFP-based GECI, K-GECO1, based on a circularly permutated RFP derived from the sea anemone Entacmaea quadricolor. We have characterized the performance of K-GECO1 in cultuRed HeLa cells, dissociated neurons, stem-cell-derived cardiomyocytes, organotypic brain slices, zebrafish spinal cord in vivo, and mouse brain in vivo. K-GECO1 is the archetype of a new lineage of GECIs based on the RFP eqFP578 scaffold. It offers high sensitivity and fast kinetics, similar or better than those of current state-of-the-art indicators, with diminished lysosomal accumulation and minimal blue-light photoactivation. Further refinements of the K-GECO1 lineage could lead to further improved variants with overall performance that exceeds that of the most highly optimized Red GECIs.

  • A genetically encoded Ca2+ indicator based on circularly permutated sea anemone Red Fluorescent Protein eqFP578
    BMC, 2018
    Co-Authors: Yi Shen, Ahmed S Abdelfattah, Hod Dana, Ronak Patel, Jamien Shea, Rosana S Molina, Bijal Rawal, Vladimir Rancic, Yufen Chang
    Abstract:

    Abstract Background Genetically encoded calcium ion (Ca2+) indicators (GECIs) are indispensable tools for measuring Ca2+ dynamics and neuronal activities in vitro and in vivo. Red Fluorescent Protein (RFP)-based GECIs have inherent advantages relative to green Fluorescent Protein-based GECIs due to the longer wavelength light used for excitation. Longer wavelength light is associated with decreased phototoxicity and deeper penetration through tissue. Red GECI can also enable multicolor visualization with blue- or cyan-excitable fluorophores. Results Here we report the development, structure, and validation of a new RFP-based GECI, K-GECO1, based on a circularly permutated RFP derived from the sea anemone Entacmaea quadricolor. We have characterized the performance of K-GECO1 in cultuRed HeLa cells, dissociated neurons, stem-cell-derived cardiomyocytes, organotypic brain slices, zebrafish spinal cord in vivo, and mouse brain in vivo. Conclusion K-GECO1 is the archetype of a new lineage of GECIs based on the RFP eqFP578 scaffold. It offers high sensitivity and fast kinetics, similar or better than those of current state-of-the-art indicators, with diminished lysosomal accumulation and minimal blue-light photoactivation. Further refinements of the K-GECO1 lineage could lead to further improved variants with overall performance that exceeds that of the most highly optimized Red GECIs

  • a genetically encoded ca2 indicator based on circularly permutated sea anemone Red Fluorescent Protein
    bioRxiv, 2017
    Co-Authors: Yi Shen, Ahmed S Abdelfattah, Hod Dana, Ronak Patel, Jamien Shea, Rosana S Molina, Bijal Rawal, Vladimir Rancic, Yufen Chang, Yingche Chen
    Abstract:

    Genetically-encoded calcium ion (Ca2+) indicators (GECIs) are indispensable tools for measuring Ca2+ dynamics and neuronal activities in vitro and in vivo. Red Fluorescent Protein (RFP)-based GECIs enable multicolor visualization with blue or cyan-excitable fluorophores and combined use with blue or cyan-excitable optogenetic actuators. Here we report the development, structure, and validation of a new Red Fluorescent Ca2+ indicator, K-GECO1, based on a circularly permutated RFP derived from the sea anemone Entacmaea quadricolor. We characterized the performance of K-GECO1 in cultuRed HeLa cells, dissociated neurons, stem cell derived cardiomyocytes, organotypic brain slices, zebrafish spinal cord in vivo, and mouse brain in vivo.

Related terms

Red Fluorescent Protein - 15 days free trial to Access Experts & Abstracts