Defining a polymethine dye for fluorescence anisotropy applications in the near-infrared spectral range.

Fluorescence anisotropy in the near-infrared (NIR) spectral range is challenging because of the lack of appropriate NIR fluorescent labels. We have evaluated polymethine fluorescent dyes to identify a leading candidate for NIR anisotropy applications. The NIR dye LS601 demonstrated low fluorescence anisotropy values (r) as a result of its relatively long fluorescent lifetime 1.3 ns. The r value of LS601 unbound and coupled to biological macromolecules was found to have a sufficient dynamic range from 0.24 to 0.37, demonstrating the feasibility of fluorescence anisotropy in the NIR. The viability of fluorescence anisotropy using a NIR label was demonstrated by characterization of dye-protein conjugates. These results open the door to a number of applications in drug discovery, fluorescence anisotropy imaging and contrast agent development.

[1]  S. Ryder,et al.  Quantitative approaches to monitor protein-nucleic acid interactions using fluorescent probes. , 2011, RNA.

[2]  B. P. Sullivan,et al.  Application of the energy gap law to the decay of charge transfer excited states, solvent effects , 1982 .

[3]  Yuexin Liu,et al.  Membrane order and molecular dynamics associated with IgE receptor cross-linking in mast cells. , 2007, Biophysical journal.

[4]  V. Tretyachenko-Ladokhina,et al.  Alexa and Oregon Green dyes as fluorescence anisotropy probes for measuring protein-protein and protein-nucleic acid interactions. , 2002, Analytical biochemistry.

[5]  S. Achilefu,et al.  Near infrared dyes as lifetime solvatochromic probes for micropolarity measurements of biological systems. , 2007, Biophysical journal.

[6]  S. Achilefu,et al.  Near-infrared fluorescence lifetime pH-sensitive probes. , 2011, Biophysical journal.

[7]  D. Taylor,et al.  Fluorescence anisotropy imaging microscopy maps calmodulin binding during cellular contraction and locomotion , 1993, The Journal of cell biology.

[8]  V. Ntziachristos,et al.  Hydrophilic Cyanine Dyes as Contrast Agents for Near-infrared Tumor Imaging: Synthesis, Photophysical Properties and Spectroscopic In vivo Characterization¶ , 2000, Photochemistry and photobiology.

[9]  D. Jameson,et al.  Fluorescence polarization/anisotropy in diagnostics and imaging. , 2010, Chemical reviews.

[10]  Samuel Achilefu,et al.  Heptamethine cyanine dyes with a robust C-C bond at the central position of the chromophore. , 2006, The Journal of organic chemistry.

[11]  Joseph R. Lakowicz,et al.  Three-photon induced fluorescence of 2,5-diphenyloxazole with a femtosecond Ti:sapphire laser , 1995 .

[12]  H. Frank,et al.  The application of the energy gap law to the S1 energies and dynamics of carotenoids , 1995 .

[13]  Sharon Bloch,et al.  Multivalent carbocyanine molecular probes: synthesis and applications. , 2005, Bioconjugate chemistry.

[14]  Ann C Horan,et al.  Use of Red-Shifted Dyes in a Fluorescence Polarization AKT Kinase Assay for Detection of Biological Activity in Natural Product Extracts , 2004, Journal of biomolecular screening.

[15]  D. Chiu,et al.  Probing rotational viscosity in synaptic vesicles. , 2011, Biophysical journal.

[16]  R. Gillies,et al.  Gold Nanorods Targeted to Delta Opioid Receptor: Plasmon-Resonant Contrast and Photothermal Agents , 2008, Molecular imaging.

[17]  Samuel Achilefu,et al.  Ratiometric Analysis of Fluorescence Lifetime for Probing Binding Sites in Albumin with Near‐Infrared Fluorescent Molecular Probes , 2007, Photochemistry and photobiology.

[18]  V. Krishnan,et al.  An empirical relationship between rotational correlation time and solvent accessible surface area , 1998, Journal of biomolecular NMR.

[19]  Yunpeng Ye,et al.  Synthesis and characterization of a macrocyclic near-infrared optical scaffold. , 2003, Journal of the American Chemical Society.

[20]  N. Murthy,et al.  Hydrocyanines: a class of fluorescent sensors that can image reactive oxygen species in cell culture, tissue, and in vivo. , 2009, Angewandte Chemie.

[21]  Joshua Jortner,et al.  The energy gap law for radiationless transitions in large molecules , 1970 .

[22]  M. Toth,et al.  Measurement of specific protease activity utilizing fluorescence polarization. , 1997, Analytical biochemistry.

[23]  I. Gryczynski,et al.  Fluorescence polarization standard for near infrared spectroscopy and microscopy. , 2008, Applied optics.

[24]  Samuel Achilefu,et al.  Fluorescence lifetime properties of near-infrared cyanine dyes in relation to their structures. , 2008, Journal of photochemistry and photobiology. A, Chemistry.

[25]  M. Irving.  Steady-state polarization from cylindrically symmetric fluorophores undergoing rapid restricted motion. , 1996, Biophysical journal.

[26]  Piotr Sawosz,et al.  Wavelength-resolved measurements of fluorescence lifetime of indocyanine green. , 2011, Journal of biomedical optics.

[27]  R. C. Benson,et al.  Fluorescence properties of indocyanine green as related to angiography. , 1978, Physics in medicine and biology.

[28]  Knut Rurack,et al.  Fluorescence quantum yields of a series of red and near-infrared dyes emitting at 600-1000 nm. , 2011, Analytical chemistry.

[29]  F. Jähnig.  Structural order of lipids and proteins in membranes: evaluation of fluorescence anisotropy data. , 1979, Proceedings of the National Academy of Sciences of the United States of America.

[30]  J. Qu,et al.  Fluorescence anisotropy of molecular rotors. , 2011, Chemphyschem : a European journal of chemical physics and physical chemistry.

[31]  Robert W. Bryant,et al.  Evaluation of Fluorescent Compound Interference in 4 Fluorescence Polarization Assays: 2 Kinases, 1 Protease, and 1 Phosphatase , 2003, Journal of biomolecular screening.

[32]  A. Kahan,et al.  Photoreactivity of a push-pull merocyanine in static electric fields: a three-state model of isomerization reactions involving conical intersections. , 2009, The journal of physical chemistry. A.

[33]  Yasuteru Urano,et al.  Development and application of a near-infrared fluorescence probe for oxidative stress based on differential reactivity of linked cyanine dyes. , 2010, Journal of the American Chemical Society.

[34]  Miki N. Newman,et al.  Utilization of Fluorescence Polarization and Time Resolved Fluorescence Resonance Energy Transfer Assay Formats for SAR Studies: Src Kinase as a Model System , 2004, Journal of biomolecular screening.

[35]  S. Ameer-Beg,et al.  Fluorescence lifetime and polarization-resolved imaging in cell biology. , 2009, Current opinion in biotechnology.

[36]  H. Maeda.  Assay of proteolytic enzymes by the fluorescence polarization technique. , 1979, Analytical biochemistry.

[37]  K. Drexhage,et al.  Stable heptamethine pyrylium dyes that absorb in the infrared , 1977 .

[38]  Ralph Weissleder,et al.  Near-infrared fluorescence: application to in vivo molecular imaging. , 2010, Current opinion in chemical biology.

[39]  J. Sportsman,et al.  A Homogeneous Fluorescence Polarization Assay Adaptable for a Range of Protein Serine/Threonine and Tyrosine Kinases , 2003, Journal of biomolecular screening.

[40]  Sergei A Eremin,et al.  Fluorescence polarization immunoassays and related methods for simple, high-throughput screening of small molecules , 2008, Analytical and bioanalytical chemistry.

[41]  Yasuteru Urano,et al.  Highly sensitive near-infrared fluorescent probes for nitric oxide and their application to isolated organs. , 2005, Journal of the American Chemical Society.

[42]  Hans Kuhn,et al.  A Quantum‐Mechanical Theory of Light Absorption of Organic Dyes and Similar Compounds , 1949 .

[43]  Benjamin G. Levine,et al.  Isomerization through conical intersections. , 2007, Annual review of physical chemistry.

[44]  R R Alfano,et al.  Time-resolved fluorescence polarization dynamics and optical imaging of Cytate: a prostate cancer receptor-targeted contrast agent. , 2008, Applied optics.

[45]  J. Rao,et al.  Fluorescence imaging in vivo: recent advances. , 2007, Current opinion in biotechnology.

[46]  J. Sportsman,et al.  Fluorescence polarization assays in signal transduction discovery. , 2003, Combinatorial chemistry & high throughput screening.

[47]  Samuel Achilefu,et al.  Rational approach to select small peptide molecular probes labeled with fluorescent cyanine dyes for in vivo optical imaging. , 2011, Biochemistry.

[48]  U. Haupts,et al.  Homogeneous fluorescence readouts for miniaturized high-throughput screening: theory and practice. , 1999, Drug discovery today.

[49]  J. Owicki,et al.  Fluorescence Polarization and Anisotropy in High Throughput Screening: Perspectives and Primer , 2000, Journal of biomolecular screening.

[50]  Indrajit Roy,et al.  Covalently dye-linked, surface-controlled, and bioconjugated organically modified silica nanoparticles as targeted probes for optical imaging. , 2008, ACS nano.

[51]  Peter Karuso,et al.  Fluorescence anisotropy assay for the traceless kinetic analysis of protein digestion. , 2008, Analytical chemistry.

[52]  Simon C Watkins,et al.  The Globular Heads of C1q Specifically Recognize Surface Blebs of Apoptotic Vascular Endothelial Cells1 , 2001, The Journal of Immunology.

[53]  A. Simeonov,et al.  Enzyme assays by fluorescence polarization in the presence of polyarginine: study of kinase, phosphatase, and protease reactions. , 2002, Analytical biochemistry.