Details of a Researcher - Hanaoka, Kenjiro

Papers - Hanaoka, Kenjiro

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A design strategy for small molecule-based targeted MRI contrast agents: Their application for detection of atherosclerotic plaques

Iwaki S., Hokamura K., Ogawa M., Takehara Y., Muramatsu Y., Yamane T., Hirabayashi K., Morimoto Y., Hagisawa K., Nakahara K., Mineno T., Terai T., Komatsu T., Ueno T., Tamura K., Adachi Y., Hirata Y., Arita M., Arai H., Umemura K., Nagano T., Hanaoka K.

Organic and Biomolecular Chemistry (Organic and Biomolecular Chemistry) 12 ( 43 ) 8611 - 8618 2014.11

Research paper (scientific journal), Accepted, ISSN 14770520

Gadolinium(III) ion (Gd3+) complexes are widely used as contrast agents in magnetic resonance imaging (MRI), and many attempts have been made to couple them to sensor moieties in order to visualize biological phenomena of interest inside the body. However, the low sensitivity of MRI has made it difficult to develop practical MRI contrast agents for in vivo imaging. We hypothesized that practical MRI contrast agents could be designed by targeting a specific biological environment, rather than a specific protein such as a receptor. To test this idea, we designed and synthesized a Gd3+-based MRI contrast agent, 2BDP3Gd, for visualizing atherosclerotic plaques by linking the Gd3+-complex to the lipophilic fluorophore BODIPY to stain lipid-rich environments. We found that 2BDP3Gd was selectively accumulated into lipid droplets of adipocytes at the cellular level. Atherosclerotic plaques in the aorta of Watanabe heritable hyperlipidemic (WHHL) rabbits were clearly visualized in T1-weighted MR images after intravenous injection of 2BDP3Gd in vivo.

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Fluorometric assay of integrin activity with a small-molecular probe that senses the binding site microenvironment

Komatsu T., Takeda A., Hanaoka K., Terai T., Ueno T., Tada Y., Nagano T., Urano Y.

Chemical Communications (Chemical Communications) 50 ( 100 ) 15894 - 15896 2014.11

Research paper (scientific journal), Accepted, ISSN 13597345

We have developed a small-molecular probe, consisting of cyclic RGD pentapeptides bearing a nitrobenzoxadiazole fluorophore at the 4′-residue, which detects integrin αVβ3 activity in terms of fluorescence intensity decrease due to quenching of the fluorophore by interaction with tyrosine-122 at the binding site of the protein. This probe appears to be suitable for a practical, high-throughput fluorescence assay to screen small-molecular modulators of integrin activity. © The Royal Society of Chemistry 2014.

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Practical fluorescence detection of acrolein in human plasma via a two-step tethering approach

Togashi M., Terai T., Kojima H., Hanaoka K., Igarashi K., Hirata Y., Urano Y., Nagano T.

Chemical Communications (Chemical Communications) 50 ( 95 ) 14946 - 14948 2014.11

Research paper (scientific journal), Accepted, ISSN 13597345

Acrolein, a cytotoxic α,β-unsaturated aldehyde and disease biomarker, was determined in plasma by means of a novel tethering strategy using Michael addition of the compound to a fluorescent dye, followed by immobilization of the product on microbeads via the aldehyde moiety. Elevation of blood acrolein was detected in mice treated with an anticancer agent cyclophosphamide, which releases acrolein upon activation. This method should be suitable for high-throughput diagnostic and clinical application. © the Partner Organisations 2014.

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Boron dipyrromethene as a fluorescent caging group for single-photon uncaging with long-wavelength visible light

Umeda N., Takahashi H., Kamiya M., Ueno T., Komatsu T., Terai T., Hanaoka K., Nagano T., Urano Y.

ACS Chemical Biology (ACS Chemical Biology) 9 ( 10 ) 2242 - 2246 2014.10

Research paper (scientific journal), Accepted, ISSN 15548929

Caged compounds are useful tools for precise spatiotemporal modulation of cell functions, but in most cases uncaging requires ultraviolet (UV) light, which is cytotoxic and has limited tissue penetration. Therefore, caged compounds that can be activated by longer-wavelength light are required. Here we describe a novel photoelimination reaction of 4-aryloxy boron dipyrromethene (BODIPY) derivatives and show that BODIPY can function as a caging group for phenol groups. We developed a novel BODIPY-caged histamine compound, which is photoactivatable with blue-green visible light to stimulate cultured HeLa cells in a spatiotemporally well-controlled manner. This caging strategy is expected to be widely applicable to develop tools for probing various cellular functions. (Chemical Equation Presented).

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Cytoprotective effects of hydrogen sulfide-releasing N-methyl-d-aspartate receptor antagonists mediated by intracellular sulfane sulfur

Marutani E., Sakaguchi M., Chen W., Sasakura K., Liu J., Xian M., Hanaoka K., Nagano T., Ichinose F.

MedChemComm (MedChemComm) 5 ( 10 ) 1577 - 1583 2014.10

Research paper (scientific journal), Accepted, ISSN 20402503

Hydrogen sulfide (H2S) exerts a host of biological effects ranging from cytotoxicity to cytoprotection. Cytotoxicity of H2S in neurodegenerative diseases may be mediated by N-methyl-d-aspartate receptor (NMDAR) activation. To exploit cytoprotective effects of H2S while minimizing its toxicity, we synthesized a series of H2S-releasing NMDAR antagonists and examined their effects against 1-methyl-4-phenylpyridinium (MPP+)-induced cell death, a cellular model of Parkinson's disease. We observed that cytoprotective effects of H2S-releasing NMDAR antagonists correlated with their ability to increase intracellular sulfane sulfur, but not H2S, levels. These studies suggest that H2S-donor compounds that increase intracellular sulfane sulfur levels are potentially useful neuroprotective agents against neurodegenerative diseases.

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Thermal or mechanical stimuli-induced photoluminescence color change of a molecular assembly composed of an amphiphilic anthracene derivative in water

Sagara Y., Komatsu T., Terai T., Ueno T., Hanaoka K., Kato T., Nagano T.

Chemistry - A European Journal (Chemistry - A European Journal) 20 ( 33 ) 10397 - 10403 2014.08

Research paper (scientific journal), Accepted, ISSN 09476539

Molecular assemblies that change photoluminescence color in response to thermal or mechanical stimulation without dissociation into the monomeric states in water are described herein. A dumbbell-shaped amphiphilic compound forms micellar molecular assemblies in water and exhibits yellow photoluminescence derived from excimer formation of the luminescent core, which contains a 2,6-diethynylanthracene moiety. Annealing of the aqueous solution induces a photoluminescence color change from yellow to green (λ em, max=558→525nm). The same photoluminescence color change is also achieved by rubbing the yellow-photoluminescence-emitting molecular assemblies adsorbed on glass substrates with cotton wool in water. The observed green photoluminescence is ascribed to micelles that are distinct from the yellow-photoluminescence-emitting micelles, on the basis of transmission electron microscopy observations, atomic force microscopy observations, and dynamic light scattering measurements. We examined the relationship between the structure of the molecular assemblies and the photophysical properties of the anthracene derivative in water before and after thermal or mechanical stimulation and concluded that thermal or mechanical stimuli-induced slight changes of the molecular-assembled structures in the micelles result in the change in the photoluminescence color from yellow to green in water. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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Rapidly rendering cells phagocytic through a cell surface display technique and concurrent Rac activation

Onuma H., Komatsu T., Arita M., Hanaoka K., Ueno T., Terai T., Nagano T., Inoue T.

Science Signaling (Science Signaling) ( 334 ) 2014.07

Research paper (scientific journal), Accepted, ISSN 19450877

Cell surfaces represent a platform through which extracellular signals that determine diverse cellular processes, including migration, division, adhesion, and phagocytosis, are transduced. Techniques to rapidly reconfigure the surface properties of living cells should thus offer the ability to harness these cellular functions. Although the molecular mechanism of phagocytosis is well characterized, the minimal molecular players that are sufficient to activate this elaborate process remain elusive.We developed and implemented a technique to present a molecule of interest at the cell surface in an inducible manner on a time scale of minutes. We simultaneously induced the cell surface display of the C2 domain of milk fat globule epidermal growth factor factor 8 (MFG-E8) and activated the intracellular small guanosine triphosphatase Rac, which stimulates actin polymerization at the cell periphery. The C2 domain binds to phosphatidylserine, a lipid exposed on the surface of apoptotic cells. By integrating the stimulation of these two processes, we converted HeLa cells into a phagocytic cell line that bound to and engulfed apoptotic human Jurkat cells. Inducing either the cell surface display of the C2 domain or activating Rac alone was not sufficient to stimulate phagocytosis, which suggests that attachment to the target cell and actin reorganization together constitute the minimal molecular events that are needed to induce phagocytosis. This cell surface display technique might be useful as part of a targeted, cell-based therapy in which unwanted cells with characteristic surface molecules could be rapidly consumed by engineered cells.

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Selective ablation of β-galactosidase-expressing cells with a rationally designed activatable photosensitizer

Ichikawa Y., Kamiya M., Obata F., Miura M., Terai T., Komatsu T., Ueno T., Hanaoka K., Nagano T., Urano Y.

Angewandte Chemie - International Edition (Angewandte Chemie - International Edition) 53 ( 26 ) 6772 - 6775 2014.06

Research paper (scientific journal), Accepted, ISSN 14337851

We have developed an activatable photosensitizer capable of specifically inducing the death of β-galactosidase-expressing cells in response to photoirradiation. By using a selenium-substituted rhodol scaffold bearing β-galactoside as a targeting substituent, we designed and synthesized HMDESeR-βGal, which has a non-phototoxic spirocyclic structure owing to the presence of the galactoside moiety. However, β-galactosidase efficiently converted HMDESeR-βGal into phototoxic HMDESeR, which exists predominantly in the open xanthene form. This structural change resulted in drastic recovery of visible-wavelength absorption and the ability to generate singlet oxygen (1O2). When HMDESeR-βGal was applied to larval Drosophila melanogaster wing disks, which express β-galactosidase only in the posterior region, photoirradiation induced cell death in the β-galactosidase-expressing region with high specificity. Bull's eye! An activatable photosensitizer capable of specifically inducing the death of β-galactosidase-expressing cells in response to light irradiation was developed. Reaction with the enzyme resulted in a dynamic structural change to the phototoxic open form (see scheme), thus enabling the specific ablation of cells of interest in living tissues. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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Primary hepatocytes from mice lacking cysteine dioxygenase show increased cysteine concentrations and higher rates of metabolism of cysteine to hydrogen sulfide and thiosulfate

Jurkowska H., Roman H., Hirschberger L., Sasakura K., Nagano T., Hanaoka K., Krijt J., Stipanuk M.

Amino Acids (Amino Acids) 46 ( 5 ) 1353 - 1365 2014.05

Research paper (scientific journal), Accepted, ISSN 09394451

The oxidation of cysteine in mammalian cells occurs by two routes: a highly regulated direct oxidation pathway in which the first step is catalyzed by cysteine dioxygenase (CDO) and by desulfhydration-oxidation pathways in which the sulfur is released in a reduced oxidation state. To assess the effect of a lack of CDO on production of hydrogen sulfide (H2S) and thiosulfate (an intermediate in the oxidation of H2S to sulfate) and to explore the roles of both cystathionine γ-lyase (CTH) and cystathionine β-synthase (CBS) in cysteine desulfhydration by liver, we investigated the metabolism of cysteine in hepatocytes isolated from Cdo1-null and wild-type mice. Hepatocytes from Cdo1-null mice produced more H2S and thiosulfate than did hepatocytes from wild-type mice. The greater flux of cysteine through the cysteine desulfhydration reactions catalyzed by CTH and CBS in hepatocytes from Cdo1-null mice appeared to be the consequence of their higher cysteine levels, which were due to the lack of CDO and hence lack of catabolism of cysteine by the cysteinesulfinate-dependent pathways. Both CBS and CTH appeared to contribute substantially to cysteine desulfhydration, with estimates of 56 % by CBS and 44 % by CTH in hepatocytes from wild-type mice, and 63 % by CBS and 37 % by CTH in hepatocytes from Cdo1-null mice. © 2014 Springer-Verlag Wien.

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Unbalanced excitability underlies offline reactivation of behaviorally activated neurons

Mizunuma M., Norimoto H., Tao K., Egawa T., Hanaoka K., Sakaguchi T., Hioki H., Kaneko T., Yamaguchi S., Nagano T., Matsuki N., Ikegaya Y.

Nature Neuroscience (Nature Neuroscience) 17 ( 4 ) 503 - 505 2014.04

Research paper (scientific journal), Accepted, ISSN 10976256

Hippocampal sharp waves (SWs)/ripples represent the reactivation of neurons involved in recently acquired memory and are crucial for memory consolidation. By labeling active cells with fluorescent protein under the control of an immediate-early gene promoter, we found that neurons that had been activated while mice explored a novel environment were preferentially reactivated during spontaneous SWs in hippocampal slices in vitro. During SWs, the reactivated neurons received strong excitatory synaptic inputs as opposed to a globally tuned network balance between excitation and inhibition. © 2014 Nature America, Inc. All rights reserved.

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Covalent attachment of mechanoresponsive luminescent micelles to glasses and polymers in aqueous conditions

Sagara Y., Komatsu T., Ueno T., Hanaoka K., Kato T., Nagano T.

Journal of the American Chemical Society (Journal of the American Chemical Society) 136 ( 11 ) 4273 - 4280 2014.03

Research paper (scientific journal), Accepted, ISSN 00027863

Covalent attachment of mechanoresponsive luminescent organic or organometallic compounds to other materials is a promising approach to develop a wide variety of mechanoresponsive luminescent materials. Here, we report covalently linkable mechanoresponsive micelles that change their photoluminescence from yellow to green in response to mechanical stimulation under aqueous conditions. These micelles are composed of a dumbbell-shaped amphiphilic pyrene derivative having amine groups at the peripheral positions of its dendrons. Using a well-established cross-linker, the micelles were covalently linked via their peripheral amine groups to the surface of glass beads, polylactic acid (PLA) beads, and living cells under aqueous conditions. Vortexing of glass beads bearing the micelles in a glass vial filled with water caused a photoluminescence color change from yellow to green. PLA beads bearing the micelles showed no change in photoluminescence color under the same conditions. We ascribe this result to the lower density and stiffness of the PLA beads, because the color of the PLA beads changed on vortexing in the presence of bare glass beads. HeLa cells and HL-60 cells bearing the micelles showed no obvious photoluminescence color change under vortexing. The structure, photophysical properties, and mechanism of photoluminescence color change of the micellar assemblies were examined. © 2014 American Chemical Society.

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Development of azo-based fluorescent probes to detect different levels of hypoxia

Piao W., Tsuda S., Tanaka Y., Maeda S., Liu F., Takahashi S., Kushida Y., Komatsu T., Ueno T., Terai T., Nakazawa T., Uchiyama M., Morokuma K., Nagano T., Hanaoka K.

Angewandte Chemie - International Edition (Angewandte Chemie - International Edition) 52 ( 49 ) 13028 - 13032 2013.12

Research paper (scientific journal), Accepted, ISSN 14337851

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A water-soluble mechanochromic luminescent pyrene derivative exhibiting recovery of the initial photoluminescence color in a high-humidity environment

Sagara Y., Komatsu T., Ueno T., Hanaoka K., Kato T., Nagano T.

Advanced Functional Materials (Advanced Functional Materials) 23 ( 42 ) 5277 - 5284 2013.11

Research paper (scientific journal), Accepted, ISSN 1616301X

Switching of the luminescence properties of molecular materials in response to mechanical stimulation is of fundamental interest and also has a range of potential applications. Herein, a water-soluble mechanochromic luminescent pyrene derivative having two hydrophilic dendrons is reported. This pyrene derivative is the first example of a mechanochromic luminescent organic compound that responds to relative humidity. Mechanical stimulation (grinding) of this pyrene derivative in the solid state results in a change of the photoluminescence from yellow to green. Subsequent exposure to water vapor induces recovery of the initial yellow photoluminescence. The color change is reversible through at least ten cycles. It is also demonstrated that this compound can be applied as a mechano-sensing material in frictional wear testing for grease, owing to its immiscibility in non-polar solvents and its non-crystalline behavior. Transmission electron microscope and atomic force microscope observations of samples prepared from dilute aqueous solutions of the pyrene derivative on suitable substrates, together with dynamic light scattering measurements for the compound in aqueous solution, indicate that this amphiphilic dumbbell-shaped molecule forms micelles in water. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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Diced electrophoresis gel assay for screening enzymes with specified activities

Komatsu T., Hanaoka K., Adibekian A., Yoshioka K., Terai T., Ueno T., Kawaguchi M., Cravatt B., Nagano T.

Journal of the American Chemical Society (Journal of the American Chemical Society) 135 ( 16 ) 6002 - 6005 2013.04

Research paper (scientific journal), Accepted, ISSN 00027863

We have established the diced electrophoresis gel (DEG) assay as a proteome-wide screening tool to identify enzymes with activities of interest using turnover-based fluorescent substrates. The method utilizes the combination of native polyacrylamide gel electrophoresis (PAGE) with a multiwell-plate- based fluorometric assay to find protein spots with the specified activity. By developing fluorescent substrates that mimic the structure of neutrophil chemoattractants, we could identify enzymes involved in metabolic inactivation of the chemoattractants. © 2013 American Chemical Society.

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Red fluorescent probe for monitoring the dynamics of cytoplasmic calcium ions

Egawa T., Hirabayashi K., Koide Y., Kobayashi C., Takahashi N., Mineno T., Terai T., Ueno T., Komatsu T., Ikegaya Y., Matsuki N., Nagano T., Hanaoka K.

Angewandte Chemie - International Edition (Angewandte Chemie - International Edition) 52 ( 14 ) 3874 - 3877 2013.04

Research paper (scientific journal), Accepted, ISSN 14337851

We see red (and yellow and green): Probe 1 was developed for the visualization of cytoplasmic Ca2+, a pivotal second messenger in many biological responses. The new probe is suitable for multicolor imaging for the simultaneous detection of metal ions or proteins and is superior to the existing red fluorescent probe Rhod-2 for the monitoring of cytoplasmic Ca2+ oscillation in cultured cells (see fluorescence images of cells with 1 (top) and Rhod-2 (bottom)). © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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TokyoGreen derivatives as specific and practical fluorescent probes for UDP-glucuronosyltransferase (UGT) 1A1

Terai T., Tomiyasu R., Ota T., Ueno T., Komatsu T., Hanaoka K., Urano Y., Nagano T.

Chemical Communications (Chemical Communications) 49 ( 30 ) 3101 - 3103 2013.03

Research paper (scientific journal), Accepted, ISSN 13597345

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Rational design of highly sensitive fluorescence probes for protease and glycosidase based on precisely controlled spirocyclization

Sakabe M., Asanuma D., Kamiya M., Iwatate R.J., Hanaoka K., Terai T., Nagano T., Urano Y.

Journal of the American Chemical Society (Journal of the American Chemical Society) 135 ( 1 ) 409 - 414 2013.01

Research paper (scientific journal), Accepted, ISSN 00027863

We have synthesized and evaluated a series of hydroxymethyl rhodamine derivatives and found an intriguing difference of intramolecular spirocyclization behavior: the acetylated derivative of hydroxymethyl rhodamine green (Ac-HMRG) exists as a closed spirocyclic structure in aqueous solution at physiological pH, whereas HMRG itself takes an open nonspirocyclic structure. Ac-HMRG is colorless and nonfluorescent, whereas HMRG is strongly fluorescent. On the basis of these findings, we have developed a general design strategy to obtain highly sensitive fluorescence probes for proteases and glycosidases, by replacing the acetyl group of Ac-HMRG with a substrate moiety of the target enzyme. Specific cleavage of the substrate moiety in the nonfluorescent probe by the target enzyme generates a strong fluorescence signal. To confirm the validity and flexibility of our strategy, we designed and synthesized fluorescence probes for leucine aminopeptidase (Leu-HMRG), fibroblast activation protein (Ac-GlyPro-HMRG), and β-galactosidase (βGal-HMRG). All of these probes were almost nonfluorescent due to the formation of spirocyclic structure, but were converted efficiently to highly fluorescent HMRG by the target enzymes. We confirmed that the probes can be used in living cells. These probes offer great practical advantages, including high sensitivity and rapid response (due to regulation of fluorescence at a single reactive site), as well as resistance to photobleaching, and are expected to be useful for a range of biological and pathological investigations. © 2012 American Chemical Society.

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Reversible off-on fluorescence probe for hypoxia and imaging of hypoxia-normoxia cycles in live cells

Takahashi S., Piao W., Matsumura Y., Komatsu T., Ueno T., Terai T., Kamachi T., Kohno M., Nagano T., Hanaoka K.

Journal of the American Chemical Society (Journal of the American Chemical Society) 134 ( 48 ) 19588 - 19591 2012.12

Research paper (scientific journal), Accepted, ISSN 00027863

We report a fully reversible off-on fluorescence probe for hypoxia. The design employs QSY-21 as a Förster resonance energy transfer (FRET) acceptor and cyanine dye Cy5 as a FRET donor, based on our finding that QSY-21 undergoes one-electron bioreduction to the radical under hypoxia, with an absorbance decrease at 660 nm. At that point, FRET can no longer occur, and the dye becomes strongly fluorescent. Upon recovery of normoxia, the radical is immediately reoxidized to QSY-21, with loss of fluorescence due to restoration of FRET. We show that this probe, RHyCy5, can monitor repeated hypoxia-normoxia cycles in live cells. © 2012 American Chemical Society.

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Design strategy for a near-infrared fluorescence probe for matrix metalloproteinase utilizing highly cell permeable boron dipyrromethene

Myochin T., Hanaoka K., Komatsu T., Terai T., Nagano T.

Journal of the American Chemical Society (Journal of the American Chemical Society) 134 ( 33 ) 13730 - 13737 2012.08

Research paper (scientific journal), Accepted, ISSN 00027863

Near-infrared (NIR) fluorescence probes are especially useful for simple and noninvasive in vivo imaging inside the body because of low autofluorescence and high tissue transparency in the NIR region compared with other wavelength regions. However, existing NIR fluorescence probes for matrix metalloproteinases (MMPs), which are tumor, atherosclerosis, and inflammation markers, have various disadvantages, especially as regards sensitivity. Here, we report a novel design strategy to obtain a NIR fluorescence probe that is rapidly internalized by free diffusion and well retained intracellularly after activation by extracellular MMPs. We designed and synthesized four candidate probes, each consisting of a cell permeable or nonpermeable NIR fluorescent dye as a Förster resonance energy transfer (FRET) donor linked to the NIR dark quencher BHQ-3 as a FRET acceptor via a MMP substrate peptide. We applied these probes for detection of the MMP activity of cultured HT-1080 cells, which express MMP2 and MT1-MMP, by fluorescence microscopy. Among them, the probe incorporating BODIPY650/665, BODIPY-MMP, clearly visualized the MMP activity as an increment of fluorescence inside the cells. We then applied this probe to a mouse xenograft tumor model prepared with HT-1080 cells. Following intratumoral injection of the probe, MMP activity could be visualized for much longer with BODIPY-MMP than with the probe containing SulfoCy5, which is cell impermeable and consequently readily washed out of the tissue. This simple design strategy should be applicable to develop a range of sensitive, rapidly responsive NIR fluorescence probes not only for MMP activity, but also for other proteases. © 2012 American Chemical Society.

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Highly activatable and environment-insensitive optical highlighters for selective spatiotemporal imaging of target proteins

Kobayashi T., Komatsu T., Kamiya M., Campos C., González-Gaitán M., Terai T., Hanaoka K., Nagano T., Urano Y.

Journal of the American Chemical Society (Journal of the American Chemical Society) 134 ( 27 ) 11153 - 11160 2012.07

Research paper (scientific journal), Accepted, ISSN 00027863

Optical highlighters are photoactivatable fluorescent molecules that exhibit pronounced changes in their spectral properties in response to irradiation with light of a specific wavelength and intensity. Here, we present a novel design strategy for a new class of caged BODIPY (4,4-difluoro-4-bora-3a, 4a-diaza-s-indacene) fluorophores, based on the use of photoremovable protecting groups (PRPGs) with high reduction potentials that serve as both a photosensitive unit and a fluorescence quencher via photoinduced electron transfer (PeT). 2,6-Dinitrobenzyl (DNB)-caged BODIPY was efficiently photoactivated, with activation ratios exceeding 600-fold in aqueous solutions. We then combined this photoactivatable fluorophore with a SNAP (mutant of O 6-alkylguanine DNA alkyltransferase) ligand to obtain a small-molecule-based optical highlighter for visualization of protein dynamics, using the well-established SNAP tag technology. As proof of concept, we demonstrate spatiotemporal imaging of the fusion protein of epidermal growth factor receptor (EGFR) with SNAP tag in living cells. We also demonstrate highlighting of cells of interest in live zebrafish embryos, using the fusion protein of histone 2A with SNAP tag. © 2012 American Chemical Society.

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