Near-infrared fluorescent proteins

Shemiakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Science, Moscow, Russia.
Nature Methods (Impact Factor: 25.95). 10/2010; 7(10):827-9. DOI: 10.1038/nmeth.1501
Source: PubMed

ABSTRACT Fluorescent proteins with emission wavelengths in the near-infrared and infrared range are in high demand for whole-body imaging techniques. Here we report near-infrared dimeric fluorescent proteins eqFP650 and eqFP670. To our knowledge, eqFP650 is the brightest fluorescent protein with emission maximum above 635 nm, and eqFP670 displays the most red-shifted emission maximum and high photostability.

Download full-text


Available from: Gary D Luker, Jul 29, 2015
  • Source
    • "A plasmid containing cDNA copy of recombinant VSV-XN2-ΔM51 genome (VSV Indiana serotype) (Lawson et al., 1995; Wollmann et al., 2010) was kindly provided by Jack Rose (Yale University). A pUC57 plasmid encoding near-infrared fluorescent protein eqFP650 was designed based on the published eqFP650 sequence (accession HQ148301) (Shcherbo et al., 2010) and was purchased from Genscript. The pUC57-eqFP650 plasmid contains a T7 promoter, a XhoI site, and a Kozak consensus sequence upstream of the eqFP650 start site (TAATACGACTCACTATAGGGAGACTCGAGCCACCATG). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Virus-encoded tumor suppressor p53 transgene expression has been successfully used in vesicular stomatitis virus (VSV) and other oncolytic viruses (OVs) to enhance their anticancer activities. However, p53 is also known to inhibit virus replication via enhanced type I interferon (IFN) antiviral responses. To examine whether p53 transgenes enhance antiviral signaling in human pancreatic ductal adenocarcinoma (PDAC) cells, we engineered novel VSV recombinants encoding human p53 or the previously described chimeric p53-CC, which contains the coiled-coil (CC) domain from breakpoint cluster region (BCR) protein and evades the dominant-negative activities of endogenously expressed mutant p53. Contrary to an expected enhancement of antiviral signaling by p53, our global analysis of gene expression in PDAC cells showed that both p53 and p53-CC dramatically inhibited type I IFN responses. Our data suggest that this occurs through p53-mediated inhibition of the NF-κB pathway. Importantly, VSV-encoded p53 or p53-CC did not inhibit antiviral signaling in non-malignant human pancreatic ductal cells, which retained their resistance to all tested VSV recombinants. To the best of our knowledge, this is the first report of p53-mediated inhibition of antiviral signaling, and it suggests that OV-encoded p53 can simultaneously produce anticancer activities while assisting, rather than inhibiting, virus replication in cancer cells. Copyright © 2015 Elsevier Inc. All rights reserved.
    Virology 05/2015; 483:126-140. DOI:10.1016/j.virol.2015.04.017 · 3.28 Impact Factor
  • Source
    • "We transduced HeyA8 cells with a secreted C-terminal fragment of Gaussia luciferase (CG). NG-CXCR4 cells also were transduced with firefly luciferase [19]. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Patients with metastatic ovarian cancer continue to have a dismal prognosis, emphasizing the need for new strategies to identify and develop new molecular targets for therapy. Chemokine CXCL12 and its receptor CXCR4 are upregulated in metastatic ovarian cancer cells and the intraperitoneal tumor microenvironment. CXCL12-CXCR4 signaling promotes multiple steps in proliferation and dissemination of ovarian cancer cells, suggesting that targeted inhibition of this pathway will limit tumor progression. To investigate CXCL12-CXCR4 signaling in ovarian cancer and establish effects of inhibiting this pathway on tumor progression and survival, we designed a Gaussia luciferase complementation imaging reporter system to detect CXCL12 binding to CXCR4 in ovarian cancer cells. In cell-based assays, we established that the complementation imaging reporter could detect CXCL12 binding to CXCR4 and quantify specific inhibition of ligand-receptor interaction. We monitored CXCL12-CXCR4 binding and inhibition in a mouse xenograft model of metastatic human ovarian cancer by imaging Gaussia luciferase complementation and assessed tumor progression with firefly luciferase. Bioluminescence imaging studies in living mice showed that treatment with AMD3100, a clinically approved inhibitor of CXCL12-CXCR4, blocked ligand-receptor binding and reduced growth of ovarian cancer cells. Treatment with AMD3100 also modestly improved overall survival of mice with metastatic ovarian cancer. The Gaussia luciferase complementation imaging reporter system will facilitate further preclinical development and optimization of CXCL12-CXCR4 targeted compounds for treatment of ovarian cancer. Our research supports clinical translation of existing CXCR4 inhibitors for molecular therapy for ovarian cancer.
    Neoplasia (New York, N.Y.) 12/2011; 13(12):1152-61. DOI:10.1593/neo.111076 · 5.40 Impact Factor
  • Source
    • "The fluorescent distribution of the brain and other organs were visualized with IVIS Live Animal Imager (Caliper Life Sciences, Hopkinton, MA, USA; courtesy of the UMMC Cancer Institute and the UMMC Laboratory Animal Facility) using an Excitation wavelength of 745 nm and an Emission wavelength of 800 nm. Radiant Efficiency is a common unit used to measure Epi-fluorescence changes in vivo (Shcherbo et al., 2010). The average Radiant Efficiency of the cerebellum ± SE was calculated by taking an ROI of the cerebellum and using the Living Image software (Caliper). "
    [Show abstract] [Hide abstract]
    ABSTRACT: S100B, a glial-secreted protein, is believed to play a major role in neurodegeneration in Alzheimer's disease, Down syndrome, traumatic brain injury, and spinocerebellar ataxia type 1 (SCA1). SCA1 is a trinucleotide repeat disorder in which the expanded polyglutamine mutation in the protein ataxin-1 primarily targets Purkinje cells of the cerebellum. Currently, the exact mechanism of S100B-mediated Purkinje cell damage in SCA1 is not clear. However, here we show that S100B may act via the activation of the receptor for advanced glycation end product (RAGE) signaling pathway, resulting in oxidative stress-mediated injury to mutant ataxin-1-expressing neurons. To combat S100B-mediated neurodegeneration, we have designed a selective thermally responsive S100B inhibitory peptide, Synb1-ELP-TRTK. Our therapeutic polypeptide was developed using three key elements: (1) the elastin-like polypeptide (ELP), a thermally responsive polypeptide, (2) the TRTK12 peptide, a known S100B inhibitory peptide, and (3) a cell-penetrating peptide, Synb1, to enhance intracellular delivery. Binding studies revealed that our peptide, Synb1-ELP-TRTK, interacts with its molecular target S100B and maintains a high S100B binding affinity as comparable with the TRTK12 peptide alone. In addition, in vitro studies revealed that Synb1-ELP-TRTK treatment reduces S100B uptake in SHSY5Y cells. Furthermore, the Synb1-ELP-TRTK peptide decreased S100B-induced oxidative damage to mutant ataxin-1-expressing neurons. To test the delivery capabilities of ELP-based therapeutic peptides to the cerebellum, we treated mice with fluorescently labeled Synb1-ELP and observed that thermal targeting enhanced peptide delivery to the cerebellum. Here, we have laid the framework for thermal-based therapeutic targeting to regions of the brain, particularly the cerebellum. Overall, our data suggest that thermal targeting of ELP-based therapeutic peptides to the cerebellum is a novel treatment strategy for cerebellar neurodegenerative disorders.
    Neuroscience 09/2011; 197:369-80. DOI:10.1016/j.neuroscience.2011.09.025 · 3.33 Impact Factor
Show more