@article{cai_zhang_fang_brenner_kweon_sun_goldberg_zhang_2024, title={Multiscale imaging of corneal endothelium damage and Rho-kinase inhibitor application in mouse models of acute ocular hypertension}, volume={15}, ISSN={["2156-7085"]}, DOI={10.1364/BOE.510432}, abstractNote={We developed a multiscale optical imaging workflow, integrating and correlating visible-light optical coherence tomography, confocal laser scanning microscopy, and single-molecule localization microscopy to investigate mouse cornea damage from the in-vivo tissue level to the nanoscopic single-molecule level. We used electron microscopy to validate the imaged nanoscopic structures. We imaged wild-type mice and mice with acute ocular hypertension and examined the effects of Rho-kinase inhibitor application. We defined four types of intercellular tight junction structures as healthy, compact, partially-distorted, and fully-distorted types by labeling the zonula occludens-1 protein in the corneal endothelial cell layer. We correlated the statistics of the four types of tight junction structures with cornea thickness and intraocular pressure. We found that the population of fully-distorted tight junctions correlated well with the level of corneal edema, and applying Rho-kinase inhibitor reduced the population of fully-distorted tight junctions under acute ocular hypertension. Together, these data point to the utility of multiscale optical imaging in revealing fundamental biology relevant to disease and therapeutics.}, number={2}, journal={BIOMEDICAL OPTICS EXPRESS}, author={Cai, Zhen and Zhang, Yang and Fang, Raymond S. and Brenner, Benjamin and Kweon, Junghun and Sun, Cheng and Goldberg, Jeffrey L. and Zhang, Hao F.}, year={2024}, month={Feb} }
 @article{brenner_xu_zhang_kweon_fang_sheibani_zhang_sun_zhang_2024, title={Quantifying nanoscopic alterations associated with mitochondrial dysfunction using three-dimensional single-molecule localization microscopy}, volume={15}, ISSN={["2156-7085"]}, DOI={10.1364/BOE.510351}, abstractNote={Mitochondrial morphology provides unique insights into their integrity and function. Among fluorescence microscopy techniques, 3D super-resolution microscopy uniquely enables the analysis of mitochondrial morphological features individually. However, there is a lack of tools to extract morphological parameters from super-resolution images of mitochondria. We report a quantitative method to extract mitochondrial morphological metrics, including volume, aspect ratio, and local protein density, from 3D single-molecule localization microscopy images, with single-mitochondrion sensitivity. We validated our approach using simulated ground-truth SMLM images of mitochondria. We further tested our morphological analysis on mitochondria that have been altered functionally and morphologically in controlled manners. This work sets the stage to quantitatively analyze mitochondrial morphological alterations associated with disease progression on an individual basis.}, number={3}, journal={BIOMEDICAL OPTICS EXPRESS}, author={Brenner, Benjamin and Xu, Fengyuanshan and Zhang, Yang and Kweon, Junghun and Fang, Raymond and Sheibani, Nader and Zhang, Sarah X. and Sun, Cheng and Zhang, Hao F.}, year={2024}, month={Mar}, pages={1571–1584} }
 @article{jiang_mcclure_mao_chen_liu_zhang_2023, title={Integrating Machine Learning and Color Chemistry: Developing a High-School Curriculum toward Real-World Problem-Solving}, volume={12}, ISSN={["1938-1328"]}, url={https://doi.org/10.1021/acs.jchemed.3c00589}, DOI={10.1021/acs.jchemed.3c00589}, abstractNote={Artificial intelligence (AI) is rapidly transforming our world, making it imperative to educate the next generation about both the potential benefits and the challenges associated with AI. This study presents a cross-disciplinary curriculum that connects AI and chemistry disciplines in the high school classroom. Particularly, we leverage machine learning (ML), an important and simple application of AI to instruct students to build an ML-based virtual pH meter for high-precision pH read-outs. We used a "codeless" and free ML neural network building software, Orange, along with a simple chemical topic of pH to show the connection between AI and chemistry for high-schoolers who might have rudimentary backgrounds in both disciplines. The goal of this curriculum is to promote student interest and drive in the analytical chemistry domain and offer insights into how the interconnection between chemistry and ML can benefit high-school students in science learning. The activity involves students using pH strips to measure the pH of various solutions with local relevancy and then building an ML neural network model to predict the pH value based on the color changes of pH strips. The integrated curriculum increased student interest in chemistry and ML and demonstrated the relevance of science to students' daily lives and global issues. This approach is transformative in developing a broad spectrum of integration topics between chemistry and ML and understanding their global impacts.}, journal={JOURNAL OF CHEMICAL EDUCATION}, author={Jiang, Shiyan and Mcclure, Jeanne and Mao, Hongjing and Chen, Jiahui and Liu, Yunshu and Zhang, Yang}, year={2023}, month={Dec} }
 @article{yeo_zhang_neely_bao_sun_zhang_2023, title={Investigating Uncertainties in Single-Molecule Localization Microscopy Using Experimentally Informed Monte Carlo Simulation}, volume={7}, ISSN={["1530-6992"]}, url={https://doi.org/10.1021/acs.nanolett.3c00852}, DOI={10.1021/acs.nanolett.3c00852}, abstractNote={Single-molecule localization microscopy (SMLM) enables the visualization of cellular nanostructures in vitro with sub-20 nm resolution. While substructures can generally be imaged with SMLM, the structural understanding of the images remains elusive. To better understand the link between SMLM images and the underlying structure, we developed a Monte Carlo (MC) simulation based on experimental imaging parameters and geometric information to generate synthetic SMLM images. We chose the nuclear pore complex (NPC), a nanosized channel on the nuclear membrane which gates nucleo-cytoplasmic transport of biomolecules, as a test geometry for testing our MC model. Using the MC model to simulate SMLM images, we first optimized our clustering algorithm to separate >106 molecular localizations of fluorescently labeled NPC proteins into hundreds of individual NPCs in each cell. We then illustrated using our MC model to generate cellular substructures with different angles of labeling to inform our structural understanding through the SMLM images obtained.}, journal={NANO LETTERS}, author={Yeo, Wei-Hong and Zhang, Yang and Neely, Amy E. and Bao, Xiaomin and Sun, Cheng and Zhang, Hao F.}, year={2023}, month={Jul} }
 @article{neely_zhang_blumensaadt_mao_brenner_sun_zhang_bao_2023, title={Nucleoporin downregulation modulates progenitor differentiation independent of nuclear pore numbers}, volume={6}, ISSN={["2399-3642"]}, url={https://doi.org/10.1038/s42003-023-05398-6}, DOI={10.1038/s42003-023-05398-6}, abstractNote={AbstractNucleoporins (NUPs) comprise nuclear pore complexes, gateways for nucleocytoplasmic transport. As primary human keratinocytes switch from the progenitor state towards differentiation, most NUPs are strongly downregulated, with NUP93 being the most downregulated NUP in this process. To determine if this NUP downregulation is accompanied by a reduction in nuclear pore numbers, we leveraged Stochastic Optical Reconstruction Microscopy. No significant changes in nuclear pore numbers were detected using three independent NUP antibodies; however, NUP reduction in other subcellular compartments such as the cytoplasm was identified. To investigate how NUP reduction influences keratinocyte differentiation, we knocked down NUP93 in keratinocytes in the progenitor-state culture condition. NUP93 knockdown diminished keratinocytes’ clonogenicity and epidermal regenerative capacity, without drastically affecting nuclear pore numbers or permeability. Using transcriptome profiling, we identified that NUP93 knockdown induces differentiation genes related to both mechanical and immune barrier functions, including the activation of known NF-κB target genes. Consistently, keratinocytes with NUP93 knockdown exhibited increased nuclear localization of the NF-κB p65/p50 transcription factors, and increased NF-κB reporter activity. Taken together, these findings highlight the gene regulatory roles contributed by differential NUP expression levels in keratinocyte differentiation, independent of nuclear pore numbers.}, number={1}, journal={COMMUNICATIONS BIOLOGY}, author={Neely, Amy E. and Zhang, Yang and Blumensaadt, Laura A. and Mao, Hongjing and Brenner, Benjamin and Sun, Cheng and Zhang, Hao F. and Bao, Xiaomin}, year={2023}, month={Oct} }
 @misc{zhang_zheng_tomassini_singh_raymo_2023, title={Photoactivatable BODIPYs for Live-Cell PALM}, volume={28}, ISSN={["1420-3049"]}, DOI={10.3390/molecules28062447}, abstractNote={Photoactivated localization microscopy (PALM) relies on fluorescence photoactivation and single-molecule localization to overcome optical diffraction and reconstruct images of biological samples with spatial resolution at the nanoscale. The implementation of this subdiffraction imaging method, however, requires fluorescent probes with photochemical and photophysical properties specifically engineered to enable the localization of single photoactivated molecules with nanometer precision. The synthetic versatility and outstanding photophysical properties of the borondipyrromethene (BODIPY) chromophore are ideally suited to satisfy these stringent requirements. Specifically, synthetic manipulations of the BODIPY scaffold can be invoked to install photolabile functional groups and photoactivate fluorescence under photochemical control. Additionally, targeting ligands can be incorporated in the resulting photoactivatable fluorophores (PAFs) to label selected subcellular components in live cells. Indeed, photoactivatable BODIPYs have already allowed the sub-diffraction imaging of diverse cellular substructures in live cells using PALM and can evolve into invaluable analytical probes for bioimaging applications.}, number={6}, journal={MOLECULES}, author={Zhang, Yang and Zheng, Yeting and Tomassini, Andrea and Singh, Ambarish Kumar and Raymo, Francisco M.}, year={2023}, month={Mar} }
 @article{zhang_zheng_tomassini_singh_raymo_2023, title={Photoactivatable Fluorophores for Bioimaging Applications}, volume={1}, url={http://dx.doi.org/10.1021/acsaom.3c00025}, DOI={10.1021/acsaom.3c00025}, abstractNote={Photoactivatable fluorophores provide the opportunity to switch fluorescence on exclusively in a selected area within a sample of interest at a precise interval of time. Such a level of spatiotemporal fluorescence control enables the implementation of imaging schemes to monitor dynamic events in real time and visualize structural features with nanometer resolution. These transformative imaging methods are contributing fundamental insights on diverse cellular processes with profound implications in biology and medicine. Current photoactivatable fluorophores, however, become emissive only after the activation event, preventing the acquisition of fluorescence images and, hence, the visualization of the sample prior to activation. We developed a family of photoactivatable fluorophores capable of interconverting between emissive states with spectrally resolved fluorescence, instead of switching from a nonemissive state to an emissive one. We demonstrated that our compounds allow the real-time monitoring of molecules diffusing across the cellular blastoderm of developing embryos as well as of polymer beads translocating along the intestinal tract of live nematodes. Additionally, they also permit the tracking of single molecules in the lysosomal compartments of live cells and the visualization of these organelles with nanometer resolution. Indeed, our photoactivatable fluorophores may evolve into invaluable analytical tools for the investigation of the fundamental factors regulating the functions and structures of cells at the molecular level.}, number={3}, journal={ACS Applied Optical Materials}, publisher={American Chemical Society (ACS)}, author={Zhang, Yang and Zheng, Yeting and Tomassini, Andrea and Singh, Ambarish Kumar and Raymo, Francisco}, year={2023}, month={Mar}, pages={640–651} }
 @article{zhang_zheng_tomassini_hayter_raymo_2023, title={Photoactivatable fluorophores for bioimaging applications}, volume={12395}, ISBN={["978-1-5106-5895-0"]}, ISSN={["1605-7422"]}, DOI={10.1117/12.2647135}, abstractNote={Single-molecule localization microscopy (SMLM) strategies based on fluorescence photoactivation permit the imaging of live cells with subdiffraction resolution and the high-throughput tracking of individual biomolecules in their interior. They rely predominantly on genetically-encoded fluorescent proteins to label live cells selectively and allow the sequential single-molecule localization of sparse populations of photoactivated fluorophores. Synthetic counterparts to these photoresponsive proteins are limited to a few remarkable examples at the present stage, mostly because of the daunting challenges in engineering biocompatible molecular constructs with appropriate photochemical and photophysical properties for live-cell SMLM. Our laboratory developed a new family of synthetic photoactivatable fluorophores specifically designed for these imaging applications. They combine a borondipyrromethene (BODIPY) fluorophore and an ortho-nitrobenzyl (ONB) photocage in a single molecular skeleton. The photoinduced ONB cleavage extends electronic delocalization to shift bathochromically the BODIPY absorption and emission bands. As a result, these photochemical transformations can be exploited to switch fluorescence on in a spectral region compatible with bioimaging applications and allow the localization of the photochemical product at the single-molecule level. Furthermore, our compounds can be delivered and operated in the interior of live cells to enable the visualization of organelles with nanometer resolution and the intracellular tracking of single photoactivated molecules.}, journal={COLLOIDAL NANOPARTICLES FOR BIOMEDICAL APPLICATIONS XVIII}, author={Zhang, Yang and Zheng, Yeting and Tomassini, Andrea and Hayter, Colin E. and Raymo, Francisco M.}, year={2023} }
 @article{song_zhang_2024, title={Spectral precision improvement with demagnifying spectral images in spectroscopic nanoscopy}, volume={41}, ISSN={["1520-8532"]}, DOI={10.1364/JOSAA.497634}, abstractNote={Spectroscopic nanoscopy (SN) has been recognized as a key functional imaging tool in cell biology and chemistry because it offers the unique capability to simultaneously obtain the spatial and spectral information for single molecules. However, it has an intrinsic issue in using the limited photon budget from single emitters divided into two imaging channels to concurrently acquire spatial and spectral images. Accordingly, this issue lowers the spatial localization and spectral precision. Although several techniques have been introduced to improve the spatial precision in SN, improving the spectral precision has been overlooked so far. Here we propose a method to improve the spectral precision by optically manipulating the width of the spectroscopic signatures using a demagnifier. We evaluate its performance using numerical simulations with systematic investigations of several underlying optimal parameters such as the demagnification factor and the integration width in the proposed configuration. We also present achievable spectral precision values with different signal and background levels. Compared to the existing SN system, the 3× demagnifier-based configuration shows an approximate 35% improvement, from 2.9 nm to 1.9 nm, in the spectral precision at the 1000 photons signal level.}, number={1}, journal={JOURNAL OF THE OPTICAL SOCIETY OF AMERICA A-OPTICS IMAGE SCIENCE AND VISION}, author={Song, Ki-Hee and Zhang, Yang}, year={2024}, month={Jan}, pages={39–44} }
 @article{liu_shahid_mao_chen_waddington_song_zhang_2023, title={Switchable and Functional Fluorophores for Multidimensional Single-Molecule Localization Microscopy}, url={https://doi.org/10.1021/cbmi.3c00045}, DOI={10.1021/cbmi.3c00045}, abstractNote={Multidimensional single-molecule localization microscopy (mSMLM) represents a paradigm shift in the realm of super-resolution microscopy techniques. It affords the simultaneous detection of single-molecule spatial locations at the nanoscale and functional information by interrogating the emission properties of switchable fluorophores. The latter is finely tuned to report its local environment through carefully manipulated laser illumination and single-molecule detection strategies. This Perspective highlights recent strides in mSMLM with a focus on fluorophore designs and their integration into mSMLM imaging systems. Particular interests are the accomplishments in simultaneous multiplexed super-resolution imaging, nanoscale polarity and hydrophobicity mapping, and single-molecule orientational imaging. Challenges and prospects in mSMLM are also discussed, which include the development of more vibrant and functional fluorescent probes, the optimization of optical implementation to judiciously utilize the photon budget, and the advancement of imaging analysis and machine learning techniques.}, journal={Chemical & Biomedical Imaging}, author={Liu, Yunshu and Shahid, Md Abul and Mao, Hongjing and Chen, Jiahui and Waddington, Michael and Song, Ki-Hee and Zhang, Yang}, year={2023}, month={Aug} }
 @article{chen_tang_wang_perez_yao_huang_zhang_king_2023, title={Techniques for navigating postsurgical adhesions: Insights into mechanisms and future directions}, volume={6}, ISSN={["2380-6761"]}, DOI={10.1002/btm2.10565}, abstractNote={AbstractPostsurgical adhesions are a common complication of surgical procedures that can lead to postoperative pain, bowel obstruction, infertility, as well as complications with future procedures. Several agents have been developed to prevent adhesion formation, such as barriers, anti‐inflammatory and fibrinolytic agents. The Food and Drug Administration (FDA) has approved the use of physical barrier agents, but they have been associated with conflicting clinical studies and controversy in the clinical utilization of anti‐adhesion barriers. In this review, we summarize the human anatomy of the peritoneum, the pathophysiology of adhesion formation, the current prevention agents, as well as the current research progress on adhesion prevention. The early cellular events starting with injured mesothelial cells and incorporating macrophage response have recently been found to be associated with adhesion formation. This may provide the key component for developing future adhesion prevention methods. The current use of physical barriers to separate tissues, such as Seprafilm®, composed of hyaluronic acid and carboxymethylcellulose, can only reduce the risk of adhesion formation at the end stage. Other anti‐inflammatory or fibrinolytic agents for preventing adhesions have only been studied within the context of current research models, which is limited by the lack of in‐vitro model systems as well as in‐depth study of in‐vivo models to evaluate the efficiency of anti‐adhesion agents. In addition, we explore emerging therapies, such as gene therapy and stem cell‐based approaches, that may offer new strategies for preventing adhesion formation. In conclusion, anti‐adhesion agents represent a promising approach for reducing the burden of adhesion‐related complications in surgical patients. Further research is needed to optimize their use and develop new therapies for this challenging clinical problem.}, journal={BIOENGINEERING & TRANSLATIONAL MEDICINE}, author={Chen, Jiahui and Tang, Xiaoqi and Wang, Ziyu and Perez, Arielle and Yao, Benjamin and Huang, Ke and Zhang, Yang and King, Martin W. W.}, year={2023}, month={Jun} }
 @article{yeo_zhang_neely_bao_sun_zhang_2022, title={Experimental Parameters-Based Monte-Carlo Simulation of Single-Molecule Localization Microscopy of Nuclear Pore Complex to Evaluate Clustering Algorithms}, url={https://doi.org/10.1101/2022.09.21.508613}, DOI={10.1101/2022.09.21.508613}, abstractNote={AbstractSingle-molecule localization microscopy (SMLM) enables the detailed visualization of nuclear pore complexes (NPC) in vitro with sub-20 nm resolution. However, it is challenging to translate the localized coordinates in SMLM images to NPC functions because different algorithms to cluster localizations as individual NPCs can be biased without ground truth for validation. We developed a Monte-Carlo simulation to generate synthetic SMLM images of NPC and used the simulated NPC images as the ground truth to evaluate the performance of six clustering algorithms. We identified HDBSCAN as the optimal clustering algorithm for NPC counting and sizing. Furthermore, we compared the clustering results between the experimental and synthetic data for NUP133, a subunit in the NPC, and found them to be in good agreement.}, author={Yeo, Wei-Hong and Zhang, Yang and Neely, Amy E. and Bao, Xiaomin and Sun, Cheng and Zhang, Hao F.}, year={2022}, month={Sep} }
 @article{zhang_wang_huang_sun_kweon_li_zhe_ying_zhang_2022, title={Minimizing Molecular Misidentification in Imaging Low-Abundance Protein Interactions Using Spectroscopic Single-Molecule Localization Microscopy}, url={https://doi.org/10.1021/acs.analchem.2c02417}, DOI={10.1021/acs.analchem.2c02417}, abstractNote={Super-resolution microscopy can capture spatiotemporal organizations of protein interactions with resolution down to 10 nm; however, the analyses of more than two proteins involving low-abundance protein are challenging because spectral crosstalk and heterogeneities of individual fluorescent labels result in molecular misidentification. Here we developed a deep learning-based imaging analysis method for spectroscopic single-molecule localization microscopy to minimize molecular misidentification in three-color super-resolution imaging. We characterized the 3-fold reduction of molecular misidentification in the new imaging method using pure samples of different photoswitchable fluorophores and visualized three distinct subcellular proteins in U2-OS cell lines. We further validated the protein counts and interactions of TOMM20, DRP1, and SUMO1 in a well-studied biological process, Staurosporine-induced apoptosis, by comparing the imaging results with Western-blot analyses of different subcellular portions.}, journal={Analytical Chemistry}, author={Zhang, Yang and Wang, Gaoxiang and Huang, Peizhou and Sun, Edison and Kweon, Junghun and Li, Qianru and Zhe, Ji and Ying, Leslie L. and Zhang, Hao F.}, year={2022}, month={Oct} }
 @article{zhang_zhang_song_lin_sun_schatz_zhang_2021, title={Investigating Single-Molecule Fluorescence Spectral Heterogeneity of Rhodamines Using High-Throughput Single-Molecule Spectroscopy}, volume={12}, url={https://doi.org/10.1021/acs.jpclett.1c00192}, DOI={10.1021/acs.jpclett.1c00192}, abstractNote={We experimentally investigated several intramolecular coordinate and environmental changes as potential causes of single-molecule fluorescence spectral heterogeneities (smFSH). We developed a high-throughput single-molecule spectroscopy method to analyze more than 5000 single-molecule emission spectra from each of 9 commonly used fluorophores with different structural rigidities and deposited on substrates with different polarities. We observed an unexpectedly high smFSH from structurally rigid Rhodamine B compared with a structurally flexible Cyanine dye-Alexa Fluor 647. Based on experimentally measured smFSH, we ruled out the system's noise uncertainty, single-molecule spectral diffusion, and environmental polarity as the primary causes of the high smFSH. We found that the rotational flexibility of N,N-dialkylated groups contributed to the smFSH. With the high smFSH observed in structurally more rigid model fluorophores, we speculated that other intramolecular coordinate and environmental changes might also contribute to the high smFSH in Rhodamines.}, number={16}, journal={The Journal of Physical Chemistry Letters}, publisher={American Chemical Society (ACS)}, author={Zhang, Yang and Zhang, Yu and Song, Ki-Hee and Lin, Wei and Sun, Cheng and Schatz, George C. and Zhang, Hao F.}, year={2021}, month={Apr}, pages={3914–3921} }
 @article{gaire_zhang_li_yu_zhang_ying_2020, title={Accelerating multicolor spectroscopic single-molecule localization microscopy using deep learning}, url={https://doi.org/10.1364/BOE.391806}, DOI={10.1364/BOE.391806}, abstractNote={Spectroscopic single-molecule localization microscopy (sSMLM) simultaneously provides spatial localization and spectral information of individual single-molecules emission, offering multicolor super-resolution imaging of multiple molecules in a single sample with the nanoscopic resolution. However, this technique is limited by the requirements of acquiring a large number of frames to reconstruct a super-resolution image. In addition, multicolor sSMLM imaging suffers from spectral cross-talk while using multiple dyes with relatively broad spectral bands that produce cross-color contamination. Here, we present a computational strategy to accelerate multicolor sSMLM imaging. Our method uses deep convolution neural networks to reconstruct high-density multicolor super-resolution images from low-density, contaminated multicolor images rendered using sSMLM datasets with much fewer frames, without compromising spatial resolution. High-quality, super-resolution images are reconstructed using up to 8-fold fewer frames than usually needed. Thus, our technique generates multicolor super-resolution images within a much shorter time, without any changes in the existing sSMLM hardware system. Two-color and three-color sSMLM experimental results demonstrate superior reconstructions of tubulin/mitochondria, peroxisome/mitochondria, and tubulin/mitochondria/peroxisome in fixed COS-7 and U2-OS cells with a significant reduction in acquisition time.}, journal={Biomedical Optics Express}, author={Gaire, Sunil Kumar and Zhang, Yang and Li, Hongyu and Yu, Ray and Zhang, Hao F. and Ying, Leslie}, year={2020}, month={May} }
 @article{wang_niekerk_zhang_du_ji_wang_baker_groeniger_raymo_mattoussi_2020, title={Compact, “Clickable” Quantum Dots Photoligated with Multifunctional Zwitterionic Polymers for Immunofluorescence and In Vivo Imaging}, volume={31}, url={https://doi.org/10.1021/acs.bioconjchem.0c00169}, DOI={10.1021/acs.bioconjchem.0c00169}, abstractNote={We detail the preparation of highly fluorescent quantum dots (QDs), surface-engineered with multifunctional polymer ligands that are compact, readily compatible with strain-promoted click conjugation, and the use of these nanocrystals in immunofluorescence and in-vivo imaging. The ligand design combines the benefits of mixed coordination (i.e., thiol and imidazole) with molecular-scale zwitterion motifs, yielding sterically-stabilized and compact QDs that present a controllable number of azide groups, for easy conjugation to biomolecules via the efficient and selective click chemistry. The polymer coating was characterized using NMR spectroscopy to extract estimates of the diffusion coefficient, hydrodynamic size and ligand density. The azide-functionalized QDs were conjugated to anti-tropomyosin receptor kinase B antibody (α-TrkB), or to the brain-derived neurotrophic factor (BDNF). These conjugates were highly effective for labeling the tropomyosin receptor kinase B (TrkB) in pyramidal neurons within cortical tissue and for monitoring the BDNF induced activation of TrkB signaling in live neuronal cells. Finally, the polymer-coated QDs were applied for in vivo imaging of Drosophila Melanogaster embryo, where the QDs remain highly fluorescent and colloidally stable, with no measurable cytotoxicity. These materials would be of great use in various imaging applications, where small size, ease of conjugation and great colloidal stability for in vivo studies are needed.}, number={5}, journal={Bioconjugate Chemistry}, publisher={American Chemical Society (ACS)}, author={Wang, Wentao and Niekerk, Erna A. and Zhang, Yang and Du, Liang and Ji, Xin and Wang, Sisi and Baker, James D. and Groeniger, Kimberly and Raymo, Françisco M. and Mattoussi, Hedi}, year={2020}, month={May}, pages={1497–1509} }
 @article{zhang_raymo_2020, title={Live-Cell Imaging at the Nanoscale with Bioconjugatable and Photoactivatable Fluorophores}, volume={31}, url={https://doi.org/10.1021/acs.bioconjchem.0c00073}, DOI={10.1021/acs.bioconjchem.0c00073}, abstractNote={Optical diffraction fundamentally limits the spatial resolution of conventional fluorescence images to length scales that are, at least, two orders of magnitude longer than the dimensions of individual molecules. As a result, the development of innovative probes and imaging schemes to overcome diffraction is very much needed to enable the investigation of the fundamental factors regulating cellular functions at the molecular level. In this context, the chemical synthesis of molecular constructs with photoactivatable fluorescence and the ability to label sub-cellular components of live cells can have transformative implications. Indeed, the fluorescence of the resulting assemblies can be activated with spatiotemporal control, even in the intracellular environment, to permit the sequential localization of individual emissive labels with precision at the nanometer level and the gradual reconstruction of images with sub-diffraction resolution. The implementation of these operating principles for sub-diffraction imaging, however, is only possible if demanding photochemical and photophysical requirements to enable photoactivation and localization as well as stringent structural requisites to allow the covalent labeling of intracellular targets in live cells are satisfied. Because of these complications, only a few synthetic photoactivatable fluorophores with appropriate performance for live-cell imaging at the nanoscale have been developed so far. Significant synthetic efforts in conjunction with spectroscopic analyses are still very much needed to advance this promising research area further and turn photoactivatable fluorophores into the imaging probes of choice for the investigation of live cells.}, number={4}, journal={Bioconjugate Chemistry}, publisher={American Chemical Society (ACS)}, author={Zhang, Yang and Raymo, Françisco M.}, year={2020}, month={Apr}, pages={1052–1062} }
 @article{zhang_raymo_2020, title={Photoactivatable fluorophores for single-molecule localization microscopy of live cells}, volume={8}, url={https://doi.org/10.1088/2050-6120/ab8c5c}, DOI={10.1088/2050-6120/ab8c5c}, abstractNote={Photochemical reactions can be designed to convert either irreversibly or reversibly a nonemissive reactant into an emissive product. The irreversible disconnection of a photocleavable group from an emissive chromophore or the reversible interconversion of a photochromic component is generally exploited to implement these operating principles for fluorescence switching. In both instances, the interplay of activating radiation, to convert the nonemissive state into the emissive species, and exciting radiation, to produce fluorescence from the latter, can be exploited to switch fluorescence on in a given area of interest at a precise interval of time. Such a level of spatiotemporal control provides the opportunity to reconstruct sub-diffraction images with resolution at the nanometer level. Indeed, closely-spaced emitters can be switched on under photochemical control at distinct intervals of time and localized independently at the single-molecule level. In combination with appropriate intracellular targeting strategies, some of these photoactivatable fluorophores can be switched and localized inside live cells to permit the visualization of sub-cellular structures with a spatial resolution that would be impossible to achieve with conventional fluorophores. As a result, photoactivatable fluorophores can become invaluable probes for the implementation of super-resolution imaging schemes aimed at the elucidation of the fundamental factors controlling cellular functions at the molecular level.}, number={3}, journal={Methods and Applications in Fluorescence}, publisher={IOP Publishing}, author={Zhang, Yang and Raymo, Françisco M}, year={2020}, month={May}, pages={032002} }
 @article{davis_zhang_yi_du_song_scott_sun_zhang_2020, title={Super-Resolution Imaging of Self-Assembled Nanocarriers Using Quantitative Spectroscopic Analysis for Cluster Extraction}, volume={36}, url={https://doi.org/10.1021/acs.langmuir.9b03149}, DOI={10.1021/acs.langmuir.9b03149}, abstractNote={Self-assembled nanocarriers have inspired a range of applications for bioimaging, diagnostics, and drug delivery. Non-invasive visualization and characterization of nanocarriers are important for understanding their structure to function relationship. However, quantitative visualization of nanocarriers in the sample's native environment remains challenging using existing technologies. Single-molecule localization microscopy (SMLM) has the potential to provide both high-resolution visualization and quantitative analysis of nanocarriers in their native environment. However, non-specific binding of fluorescent probes used in SMLM can introduce artifacts, which impose challenges in quantitative analysis of SMLM images. We showed the feasibility of using spectroscopic point accumulation for imaging in nanoscale topography (sPAINT) to visualize self-assembled polymersomes (PS) with molecular specificity. Furthermore, we analyzed the unique spectral signatures of Nile Red (NR) molecules bound to the PS to reject artifacts from non-specific NR bindings. We further developed quantitative spectroscopic analysis for cluster extraction (qSPACE) to increase the localization density by 4-fold compared to sPAINT; thus, reducing variations in PS size measurements to less than 5%. Finally, using qSPACE we quantitatively imaged PS at various concentrations in aqueous solutions with ~20-nm localization precision and 97% reduction in sample misidentification relative to conventional SMLM.}, number={9}, journal={Langmuir}, publisher={American Chemical Society (ACS)}, author={Davis, Janel L. and Zhang, Yang and Yi, Sijia and Du, Fanfan and Song, Ki-Hee and Scott, Evan A. and Sun, Cheng and Zhang, Hao F.}, year={2020}, month={Mar}, pages={2291–2299} }
 @article{sansalone_zhang_mazza_davis_song_captain_zhang_raymo_2019, title={High-Throughput Single-Molecule Spectroscopy Resolves the Conformational Isomers of BODIPY Chromophores}, volume={10}, url={https://doi.org/10.1021/acs.jpclett.9b02250}, DOI={10.1021/acs.jpclett.9b02250}, abstractNote={A borondipyrromethene (BODIPY) chromophore is connected to a benzoxazole, benzothiazole or nitrobenzothiazole heterocycle through an olefinic bridge with trans configuration. Rotation about the two [C-C] bonds flanking the olefinic bridge occurs with fast kinetics in solution, leading to the equilibration of four conformational isomers for each compound. Ensemble spectroscopic measurements in solutions fail to distinguish the co-existing isomers. They reveal instead averaged absorption and emission bands with dependence of the latter on the excitation wavelength. Using high-throughput single-molecule spectroscopy, two main populations of single molecules with distinct spectral centroids are observed for each compound on glass substrates. Computational analyses suggest the two populations of molecules to be conformational isomers with antiperiplanar and periplanar arrangements of the BODIPY chromophores about its [C-C] bond to the olefinic bridge. Thus, statistical analysis of multiple single-molecule emission spectra can discriminate stereoisomers that would otherwise be impossible to distinguish by ensemble measurements alone.}, number={21}, journal={The Journal of Physical Chemistry Letters}, publisher={American Chemical Society (ACS)}, author={Sansalone, Lorenzo and Zhang, Yang and Mazza, Mercedes M. A. and Davis, Janel L. and Song, Ki-Hee and Captain, Burjor and Zhang, Hao F. and Raymo, Françisco M.}, year={2019}, month={Nov}, pages={6807–6812} }
 @article{zhang_zhang_ying_sun_zhang_2019, title={Machine-learning based spectral classification for spectroscopic single-molecule localization microscopy}, url={https://doi.org/10.1364/OL.44.005864}, DOI={10.1364/OL.44.005864}, abstractNote={Spectroscopic single-molecule localization microscopy (sSMLM) simultaneously captures the spatial locations and emission spectra of single molecular emissions and enables simultaneous multicolor super-resolution imaging. Existing sSMLM relies on extracting spectral signatures, such as weighted spectral centroids, to distinguish different molecular labels. However, the rich information carried by the complete spectral profiles is not fully utilized; thus, the misclassification rate between molecular labels can be high at low spectral analysis photon budget. We developed a machine learning (ML)-based method to analyze the full spectral profiles of each molecular emission and reduce the misclassification rate. We experimentally validated our method by imaging immunofluorescently labeled COS-7 cells using two far-red dyes typically used in sSMLM (AF647 and CF660) to resolve mitochondria and microtubules, respectively. We showed that the ML method achieved 10-fold reduction in misclassification and two-fold improvement in spectral data utilization comparing with the existing spectral centroid method.}, journal={Optics Letters}, author={Zhang, Zheyuan and Zhang, Yang and Ying, Leslie and Sun, Cheng and Zhang, Hao F.}, year={2019}, month={Dec} }
 @article{zhang_song_dong_davis_shao_sun_zhang_2019, title={Multicolor super-resolution imaging using spectroscopic single-molecule localization microscopy with optimal spectral dispersion}, url={https://doi.org/10.1364/AO.58.002248}, DOI={10.1364/AO.58.002248}, abstractNote={We developed transmission diffraction grating-based spectroscopic single-molecule localization microscopy (sSMLM) to collect the spatial and spectral information of single-molecule blinking events concurrently. We characterized the spectral heterogeneities of multiple far-red emitting dyes in a high-throughput manner using sSMLM. We also investigated the influence of spectral dispersion on the single-molecule identification performance of fluorophores with large spectral overlapping. The careful tuning of spectral dispersion in grating-based sSMLM permitted simultaneous three-color super-resolution imaging in fixed cells with a single objective lens at a relatively low photon budget. Our sSMLM has a compact optical design and can be integrated with conventional localization microscopy to provide add-on spectroscopic analysis capability.}, journal={Applied Optics}, author={Zhang, Yang and Song, Ki-Hee and Dong, Biqin and Davis, Janel L. and Shao, Guangbin and Sun, Cheng and Zhang, Hao F.}, year={2019}, month={Mar} }
 @article{song_zhang_wang_sun_zhang_2019, title={Three-dimensional biplane spectroscopic single-molecule localization microscopy}, url={https://doi.org/10.1364/OPTICA.6.000709}, DOI={10.1364/OPTICA.6.000709}, abstractNote={Spectroscopic single-molecule localization microscopy (sSMLM) captures the full emission spectra of individual molecules while simultaneously localizing their spatial locations at a precision greatly exceeding the optical diffraction limit. To achieve this, sSMLM uses a dispersive optical component to separate the emitted photons into dedicated spatial and spectral imaging channels for simultaneous acquisition. While adding a cylindrical lens in the spatial imaging channel enabled three-dimensional (3D) imaging in sSMLM, the inherent astigmatism leads to technical hurdles in spectral calibration and nonuniform lateral resolution at different depths. We found that implementing the biplane method based on the already established spatial and spectral imaging channels offers a much more attractive solution for 3D sSMLM. It allows for more efficient use of the limited photon budget and provides homogeneous lateral resolution compared with the astigmatism-based method using a cylindrical lens. Here we report 3D biplane sSMLM and demonstrate its multi-color 3D imaging capability by imaging microtubules and mitochondria in fixed COS-7 cells immunostained with Alexa Fluor 647 and CF 660C dyes, respectively. We showed a lateral localization precision of 20 nm at an average photon count of 550, a spectral precision of 4 nm at an average photon count of 1250, and an axial localization resolution of 50 nm.}, journal={Optica}, author={Song, Ki-Hee and Zhang, Yang and Wang, Gaoxiang and Sun, Cheng and Zhang, Hao F.}, year={2019}, month={Jun} }
 @article{song_zhang_wang_sun_zhang_2019, title={Three-dimensional biplane spectroscopic single-molecule localization microscopy: erratum}, url={https://doi.org/10.1364/OPTICA.6.001374}, DOI={10.1364/OPTICA.6.001374}, journal={Optica}, author={Song, Ki-Hee and Zhang, Yang and Wang, Gaoxiang and Sun, Cheng and Zhang, Hao F.}, year={2019}, month={Oct} }
 @article{sansalone_tang_garcia-amorós_zhang_nonell_baker_captain_raymo_2018, title={A Photoactivatable Far-Red/Near-Infrared BODIPY To Monitor Cellular Dynamics in Vivo}, volume={3}, url={https://doi.org/10.1021/acssensors.8b00262}, DOI={10.1021/acssensors.8b00262}, abstractNote={A mechanism to photoactivate far-red/near-infrared fluorescence with infinite contrast and under mild visible illumination was designed around the photophysical properties of borondipyrromethene (BODIPY) dyes and the photochemical behavior of oxazine heterocycles. Specifically, the photoinduced and irreversible cleavage of an oxazine ring with a laser line at 405 nm extends the electronic conjugation of a BODIPY chromophore over a 3 H-indole auxochrome with a 2-(4-methoxyphenyl)ethenyl substituent in position 5. This structural transformation shifts bathochromically the main absorption band of the BODIPY component to allow the selective excitation of the photochemical product with a laser line of 633 nm and produce fluorescence between 600 and 850 nm. This combination of activation, excitation, and emission wavelengths permits the visualization of the cellular blastoderm of developing Drosophila melanogaster embryos with optimal contrast and essentially no autofluorescence from the biological specimen. Furthermore, the sequential acquisition of images, after the photoactivation event, enables the tracking of individual cells within the embryos in real time. Thus, our structural design and operating principles for the photoactivation of far-red/near-infrared fluorescence can evolve into invaluable probes to monitor cellular dynamics in vivo.}, number={7}, journal={ACS Sensors}, publisher={American Chemical Society (ACS)}, author={Sansalone, Lorenzo and Tang, Sicheng and Garcia-Amorós, Jaume and Zhang, Yang and Nonell, Santi and Baker, James D. and Captain, Burjor and Raymo, Françisco M.}, year={2018}, month={Jul}, pages={1347–1353} }
 @article{zhang_song_tang_ravelo_cusido_sun_zhang_raymo_2018, title={Far-Red Photoactivatable BODIPYs for the Super-Resolution Imaging of Live Cells}, volume={140}, url={https://doi.org/10.1021/jacs.8b09099}, DOI={10.1021/jacs.8b09099}, abstractNote={The photoinduced disconnection of an oxazine heterocycle from a borondipyrromethene (BODIPY) chromophore activates bright far-red fluorescence. The high brightness of the product and the lack of autofluorescence in this spectral region allow its detection at the single-molecule level within the organelles of live cells. Indeed, these photoactivatable fluorophores localize in lysosomal compartments and remain covalently immobilized within these organelles. The suppression of diffusion allows the reiterative reconstruction of subdiffraction images and the visualization of the labeled organelles with excellent localization precision. Thus, the combination of photochemical, photophysical and structural properties designed into our fluorophores enable the visualization of live cells with a spatial resolution that is inaccessible to conventional fluorescence imaging.}, number={40}, journal={Journal of the American Chemical Society}, publisher={American Chemical Society (ACS)}, author={Zhang, Yang and Song, Ki-Hee and Tang, Sicheng and Ravelo, Laura and Cusido, Janet and Sun, Cheng and Zhang, Hao F. and Raymo, Françisco M.}, year={2018}, month={Oct}, pages={12741–12745} }
 @article{zhang_tang_thapaliya_sansalone_raymo_2018, title={Fluorescence activation with switchable oxazines}, url={https://doi.org/10.1039/C8CC03094D}, DOI={10.1039/C8CC03094D}, abstractNote={Activatable fluorophores allow the spatiotemporal control of fluorescence required to acquire subdiffraction images, highlight cancer cells and monitor dynamic events}, journal={Chemical Communications}, publisher={Royal Society of Chemistry (RSC)}, author={Zhang, Yang and Tang, Sicheng and Thapaliya, Ek Raj and Sansalone, Lorenzo and Raymo, Françisco M.}, year={2018} }
 @article{tang_zhang_dhakal_ravelo_anderson_collins_raymo_2018, title={Photochemical Barcodes}, volume={140}, url={https://doi.org/10.1021/jacs.8b00887}, DOI={10.1021/jacs.8b00887}, abstractNote={A photochemical strategy to encode fluorescence signals in vivo with spatial control was designed around the unique properties of a photoactivatable borondipyrromethene (BODIPY). The photoinduced disconnection of two oxazines, flanking a single BODIPY, in two consecutive steps produces a mixture of three emissive molecules with resolved fluorescence inside polymer beads. The relative amounts and emission intensities of the three fluorophores can be regulated precisely in each bead by adjusting the dose of activating photons to mark individual particles with distinct codes of fluorescence signals. The visible wavelengths and mild illumination sufficient to induce these transformations permit the photochemical barcoding of beads also in living nematodes. Different regions of the same animal can be labeled with distinct barcodes to allow the monitoring of their dynamics for long times with no toxic effects. Thus, our photochemical strategy for the generation of fluorescence barcodes can produce multiple and distinguishable labels in the same biological sample to enable the spatiotemporal tracking of, otherwise indistinguishable, targets.}, number={13}, journal={Journal of the American Chemical Society}, publisher={American Chemical Society (ACS)}, author={Tang, Sicheng and Zhang, Yang and Dhakal, Pravat and Ravelo, Laura and Anderson, Cheyenne L. and Collins, Kevin M. and Raymo, Françisco M.}, year={2018}, month={Apr}, pages={4485–4488} }
 @article{liu_zhang_baker_raymo_2017, title={A photoactivatable light tracer}, url={https://doi.org/10.1039/C7TC05049F}, DOI={10.1039/C7TC05049F}, abstractNote={The photoinduced interconversion of the two emissive states of a photoactivatable fluorophore permits the quantitative mapping of the electromagnetic radiation propagating through microscaled objects in three dimensions.}, journal={Journal of Materials Chemistry C}, publisher={Royal Society of Chemistry (RSC)}, author={Liu, Xiaoming and Zhang, Yang and Baker, James D. and Raymo, Françisco M.}, year={2017} }
 @article{thapaliya_zhang_dhakal_brown_wilson_collins_raymo_2017, title={Bioimaging with Macromolecular Probes Incorporating Multiple BODIPY Fluorophores}, volume={28}, DOI={10.1021/acs.bioconjchem.7b00166}, abstractNote={Seven macromolecular constructs incorporating multiple borondipyrromethene (BODIPY) fluorophores along a common poly(methacrylate) backbone with decyl and oligo(ethylene glycol) side chains were synthesized. The hydrophilic oligo(ethylene glycol) components impose solubility in aqueous environment on the overall assembly. The hydrophobic decyl chains effectively insulate the fluorophores from each other to prevent detrimental interchromophoric interactions and preserve their photophysical properties. As a result, the brightness of these multicomponent assemblies is approximately three times greater than that of a model BODIPY monomer. Such a high brightness level is maintained even after injection of the macromolecular probes in living nematodes, allowing their visualization with a significant improvement in signal-to-noise ratio, relative to the model monomer, and no cytotoxic or behavioral effects. The covalent scaffold of these macromolecular constructs also permits their subsequent conjugation to secondary antibodies. The covalent attachment of polymer and biomolecule does not hinder the targeting ability of the latter and the resulting bioconjugates can be exploited to stain the tubulin structure of model cells to enable their visualization with optimal signal-to-noise ratios. These results demonstrate that this particular structural design for the incorporation of multiple chromophores within the same covalent construct is a viable one to preserve the photophysical properties of the emissive species and enable the assembly of bioimaging probes with enhanced brightness.}, number={5}, journal={Bioconjugate Chemistry}, publisher={American Chemical Society (ACS)}, author={Thapaliya, Ek Raj and Zhang, Yang and Dhakal, Pravat and Brown, Adrienne S. and Wilson, James N. and Collins, Kevin M. and Raymo, Françisco M.}, year={2017}, month={May}, pages={1519–1528} }
 @article{ma_tao_zhang_li_raymo_cui_2017, title={Detection of nitroaromatic explosives by a 3D hyperbranched σ–π conjugated polymer based on a POSS scaffold}, DOI={10.1039/c7ta04351a}, abstractNote={A three-dimensional hyperbranched polymer (3D-HP) with σ–π conjugated PDMPS (poly(dichloromethylphenylsilane)s) units covalently bonded to a polyhedral oligosilsesquioxane (POSS) scaffold was prepared by a one-step “thiol–ene click chemistry” reaction.}, journal={J. Mater. Chem. A}, publisher={Royal Society of Chemistry (RSC)}, author={Ma, Xiaoshuang and Tao, Furong and Zhang, Yang and Li, Tianduo and Raymo, Françisco M. and Cui, Yuezhi}, year={2017} }
 @article{li_zhang_qiao_tao_li_ding_raymo_cui_2017, title={Facile fabrication of AIE/AIEE-active fluorescent nanoparticles based on barbituric for cell imaging applications}, volume={7}, DOI={10.1039/c7ra03956e}, abstractNote={Compounds 1–4 were synthesized, and the optical properties and mechanisms were investigated. Compound 4 was applied to the cell imaging.}, number={48}, journal={RSC Adv.}, publisher={Royal Society of Chemistry (RSC)}, author={Li, Kai and Zhang, Yang and Qiao, Bing and Tao, Furong and Li, Tianduo and Ding, Yunqiao and Raymo, Françisco M. and Cui, Yuezhi}, year={2017}, pages={30229–30241} }
 @article{thapaliya_zhang_raymo_2017, title={Fluorescence patterning with mild illumination in polymer films of photocleavable oxazines}, volume={5}, DOI={10.1039/c6tc05532j}, abstractNote={The photoinduced cleavage of oxazine heterocycles, connected to macromolecules spin coated on appropriate substrates, occurs efficiently and irreversibly to quench the emission of a fluorescent dopant and allow the imprinting of patterns.}, number={5}, journal={J. Mater. Chem. C}, publisher={Royal Society of Chemistry (RSC)}, author={Thapaliya, Ek Raj and Zhang, Yang and Raymo, Françisco M.}, year={2017}, pages={1179–1183} }
 @inproceedings{tang_zhang_thapaliya_brown_wilson_raymo_2017, title={Highlighting cancer cells with macromolecular probes}, DOI={10.1117/12.2261495}, abstractNote={Conventional fluorophore–ligand constructs for the detection of cancer cells generally produce relatively weak signals with modest contrast. The inherently low brightness accessible per biding event with the pairing of a single organic fluorophore to a single ligand as well as the contribution of unbound probes to background fluorescence are mainly responsible for these limitations. Our laboratories identified a viable structural design to improve both brightness and contrast. It is based on the integration of activatable fluorophores and targeting ligands within the same macromolecular construct. The chromophoric components are engineered to emit bright fluorescence exclusively in acidic environments. The targeting agents are designed to bind complementary receptors overexpressed on the surface of cancer cells and allow internalization of the macromolecules into acidic organelles. As a result of these properties, our macromolecular probes switch their intense emission on exclusively in the intracellular space of target cells with minimal background fluorescence from the extracellular matrix. In fact, these operating principles translate into a 170-fold enhancement in brightness, relative to equivalent but isolated chromophoric components, and a 3-fold increase in contrast, relative to model but non-activatable fluorophores. Thus, our macromolecular probes might ultimately evolve into valuable analytical tools to highlight cancer cells with optimal signal-to-noise ratios in a diversity of biomedical applications.}, booktitle={Colloidal Nanoparticles for Biomedical Applications XII}, publisher={SPIE}, author={Tang, Sicheng and Zhang, Yang and Thapaliya, Ek Raj and Brown, Adrienne S. and Wilson, James N. and Raymo, Françisco M.}, editor={Osiński, Marek and Parak, Wolfgang J. and Liang, Xing-JieEditors}, year={2017}, month={Feb} }
 @inproceedings{thapaliya_fowley_callan_tang_zhang_callan_raymo_2017, title={Probing the intracellular fate of supramolecular nanocarriers and their cargo with FRET schemes}, DOI={10.1117/12.2269514}, abstractNote={We designed a strategy to monitor self-assembling supramolecular nanocarriers and their cargo simultaneously in the intracellular space with fluorescence measurements. It is based on Fӧrster resonance energy transfer (FRET) between complementary chromophores covalently integrated in the macromolecular backbone of amphiphilic polymers and/or noncovalently encapsulated in supramolecular assemblies of the amphiphilic components. Indeed, these polymers assemble into a micelles in aqueous phase to bring energy donors and acceptors in close proximity and allow energy transfer. The resulting supramolecular assemblies maintain their integrity after travelling into the intracellular space and do not lose their molecular guests in the process. Furthermore, this mechanism can also be exploited to probe the fate of complementary nanoparticles introduced within cells in consecutive incubation steps. Efficient energy transfer occurs in the intracellular space after the sequential incubation of nanocarriers incorporating donors first and then nanoparticles containing acceptors or vice versa. The two sets of nanostructured assemblies ultimately co-localize in the cell interior to bring donors and acceptors together and enable energy transfer. Thus, this protocol is particularly valuable to monitor the transport properties of supramolecular nanocarriers inside living cells and can eventually contribute to the fundamental understating of the ability of these promising vehicles to deliver contrast agents and/or drugs intracellularly in view of possible diagnostics and/or therapeutic applications.}, booktitle={Colloidal Nanoparticles for Biomedical Applications XII}, publisher={SPIE}, author={Thapaliya, Ek Raj and Fowley, Colin and Callan, Bridgeen and Tang, Sicheng and Zhang, Yang and Callan, John F. and Raymo, Françisco M.}, editor={Osiński, Marek and Parak, Wolfgang J. and Liang, Xing-JieEditors}, year={2017}, month={Feb} }
 @inproceedings{zhang_tang_sansalone_thapaliya_baker_raymo_2017, title={Supramolecular delivery of photoactivatable fluorophores in developing embryos}, DOI={10.1117/12.2261721}, abstractNote={The identification of noninvasive strategies to monitor dynamics within living organisms in real time is essential to elucidate the fundamental factors governing a diversity of biological processes. This study demonstrates that the supramolecular delivery of photoactivatable fluorophores in Drosophila melanogaster embryos allows the real-time tracking of translocating molecules. The designed photoactivatable fluorophores switch from an emissive reactant to an emissive product with spectrally-resolved fluorescence, under moderate blue-light irradiation conditions. These hydrophobic fluorescent probes can be encapsulated within supramolecular hosts and delivered to the cellular blastoderm of the embryos. Thus, the combination of supramolecular delivery and fluorescence photoactivation translates into a noninvasive method to monitor dynamics in vivo and can evolve into a general chemical tool to track motion in biological specimens.}, booktitle={Colloidal Nanoparticles for Biomedical Applications XII}, publisher={SPIE}, author={Zhang, Yang and Tang, Sicheng and Sansalone, Lorenzo and Thapaliya, Ek Raj and Baker, James D. and Raymo, Françisco M.}, editor={Osiński, Marek and Parak, Wolfgang J. and Liang, Xing-JieEditors}, year={2017}, month={Feb} }
 @article{zhang_tang_sansalone_baker_raymo_2016, title={A Photoswitchable Fluorophore for the Real-Time Monitoring of Dynamic Events in Living Organisms}, volume={22}, DOI={10.1002/chem.201603545}, abstractNote={AbstractThis study reports the synthesis of a photoactivatable fluorophore with optimal photochemical and photophysical properties for the real‐time tracking of motion in vivo. The photoactivation mechanism designed into this particular compound permits the conversion of an emissive reactant into an emissive product with resolved fluorescence, under mild illumination conditions that are impossible to replicate with conventional switching schemes based on bleaching. Indeed, the supramolecular delivery of these photoswitchable probes into the cellular blastoderm of Drosophila melanogaster embryos allows the real‐time visualization of translocating molecules with no detrimental effects on the developing organisms. Thus, this innovative mechanism for fluorescence photoactivation can evolve into a general chemical tool to monitor dynamic processes in living biological specimens.}, number={42}, journal={Chemistry - A European Journal}, publisher={Wiley-Blackwell}, author={Zhang, Yang and Tang, Sicheng and Sansalone, Lorenzo and Baker, James D. and Raymo, Françisco M.}, year={2016}, month={Aug}, pages={15027–15034} }
 @article{zhang_captain_raymo_2016, title={A pH-Gated Photocage}, volume={5}, DOI={10.1002/adom.201600231}, abstractNote={Compounds 1 and 2 incorporate carboxyl and hydroxyl substituents in positions 2,3 and 1,2, respectively, of a naphthalene scaffold. Both molecules are commercially available and can be converted into dioxinones in a single synthetic step ( Figure 1 ). Specifi cally, their treatment with an excess of thionyl chloride and either acetone or acetophenone in dimethoxyethane, under the assistance of 4-dimethylaminopyridine, produces 3–6. Electrospray ionization mass spectra (ESIMS) of the products reveal peaks at m / z values for the corresponding molecular ions. Similarly, 1 H and 13 C nuclear magnetic resonance (NMR) spectra show resonances consistent with the structures of 3–6 and support the formation of dioxinone rings. Furthermore, high-performance liquid chromatography (HPLC) confi rms the purity of all compounds (Figures S1–S4, Supporting Information). The absorption spectra of 3–6 in methanol show intense bands between 300 and 370 nm. Illumination of the samples within this range of wavelengths for up to 10 min (2.25 mW cm −2 ), however, causes signifi cant spectral changes only for 4 and 6 ( Figure 2 and Figures S6–S8 (Supporting Information)). In agreement with literature precedents, [ 10–13 ]}, journal={Advanced Optical Materials}, publisher={Wiley-Blackwell}, author={Zhang, Yang and Captain, Burjor and Raymo, Françisco M.}, year={2016}, month={May} }
 @article{tang_zhang_thapaliya_brown_wilson_raymo_2016, title={Highlighting Cancer Cells with Halochromic Switches}, volume={2}, DOI={10.1021/acssensors.6b00592}, abstractNote={Halochromic coumarin-oxazine prefluorophores and targeting folate ligands can be connected covalently to the side chains of amphiphilic polymers. The resulting macromolecular constructs assemble into nanoparticles in aqueous environments. The prefluorophores do not produce any detectable fluorescence at neutral pH, but are converted into fluorophores with intense visible emission at acidic pH. Protonation opens the oxazine heterocycle to shift bathochromically the coumarin absorption and activate fluorescence with a brightness per nanoparticle approaching 5 × 105 M-1 cm-1. This value translates into a 170-fold enhancement relative to the isolated fluorophores dissolved in organic solvent. The folate ligands direct these multicomponent constructs into acidic intracellular compartments of folate-positive cells, where the prefluorophores switch to the corresponding fluorophores and produce fluorescence. The pH-induced activation of the signaling units ensures negligible background fluorescence from the extracellular matrix, which instead limits considerably the contrast accessible with model systems incorporating conventional nonactivatable fluorophores. Furthermore, no intracellular fluorescence can be detected when the very same measurements are performed with folate-negative cells. Nonetheless, control experiments demonstrate that the covalent connection of the prefluorophores to the polymer backbone of the amphiphilic constructs is essential to ensure selectivity. Model systems with prefluorophores noncovalently encapsulated cannot discriminate folate-positive from -negative cells. Thus, our structural design for the covalent integration of activatable signaling units and targeting ligands within the same nanostructured assembly together with the photophysical properties engineered into the emissive components offer the opportunity to highlight cancer cells selectively with high brightness and optimal contrast.}, number={1}, journal={ACS Sensors}, publisher={American Chemical Society (ACS)}, author={Tang, Sicheng and Zhang, Yang and Thapaliya, Ek Raj and Brown, Adrienne S. and Wilson, James N. and Raymo, Françisco M.}, year={2016}, pages={92–101} }
 @article{sansalone_tang_zhang_thapaliya_raymo_garcia-amorós_2016, title={Semiconductor Quantum Dots with Photoresponsive Ligands}, DOI={10.1007/978-3-319-51192-4_2}, abstractNote={Photochromic or photocaged ligands can be anchored to the outer shell of semiconductor quantum dots in order to control the photophysical properties of these inorganic nanocrystals with optical stimulations. One of the two interconvertible states of the photoresponsive ligands can be designed to accept either an electron or energy from the excited quantum dots and quench their luminescence. Under these conditions, the reversible transformations of photochromic ligands or the irreversible cleavage of photocaged counterparts translates into the possibility to switch luminescence with external control. As an alternative to regulating the photophysics of a quantum dot via the photochemistry of its ligands, the photochemistry of the latter can be controlled by relying on the photophysics of the former. The transfer of excitation energy from a quantum dot to a photocaged ligand populates the excited state of the species adsorbed on the nanocrystal to induce a photochemical reaction. This mechanism, in conjunction with the large two-photon absorption cross section of quantum dots, can be exploited to release nitric oxide or to generate singlet oxygen under near-infrared irradiation. Thus, the combination of semiconductor quantum dots and photoresponsive ligands offers the opportunity to assemble nanostructured constructs with specific functions on the basis of electron or energy transfer processes. The photoswitchable luminescence and ability to photoinduce the release of reactive chemicals, associated with the resulting systems, can be particularly valuable in biomedical research and can, ultimately, lead to the realization of imaging probes for diagnostic applications as well as to therapeutic agents for the treatment of cancer.}, journal={Photoactive Semiconductor Nanocrystal Quantum Dots}, publisher={Springer International Publishing}, author={Sansalone, Lorenzo and Tang, Sicheng and Zhang, Yang and Thapaliya, Ek Raj and Raymo, Françisco M. and Garcia-Amorós, Jaume}, year={2016}, month={Oct}, pages={31–60} }
 @article{zhang_thapaliya_tang_baker_raymo_2016, title={Supramolecular delivery of fluorescent probes in developing embryos}, volume={6}, DOI={10.1039/c6ra15368b}, abstractNote={Self-assembling nanocarriers of amphiphilic polymers encapsulate hydrophobic fluorophores in their hydrophobic interior and, upon injection in Drosophila melanogaster embryos, release their cargo into the cellular blastoderm.}, number={76}, journal={RSC Adv.}, publisher={Royal Society of Chemistry (RSC)}, author={Zhang, Yang and Thapaliya, Ek Raj and Tang, Sicheng and Baker, James D. and Raymo, Françisco M.}, year={2016}, pages={72756–72760} }
 @inproceedings{zhang_tang_thapaliya_raymo_2016, title={Supramolecular nanocarriers with photoresponsive cargo}, DOI={10.1117/12.2207926}, abstractNote={The covalent integration of fluorescent and photoswitchable components within the same molecular skeleton can be exploited to activate fluorescence under optical control. Specifically, a photoswitchable oxazine heterocycle can be connected to either a coumarin or a borondipyrromethene fluorophore. Illumination of the resulting molecular dyads at an appropriate activation wavelength either opens the heterocycle reversibly or cleaves it irreversibly, depending on the relative positions of its methylene and nitro substituents. These photochemical transformations shift bathochromically the main absorption band of the fluorophore and allow its selective excitation at a given wavelength. These hydrophobic molecular dyads can be entrapped within the hydrophobic interior of self-assembling nanoparticles of amphiphilic polymer. The supramolecular envelope around the switchable compounds enables their transfer into aqueous environments and their operation under these conditions with minimal influence on their photochemical and photophysical properties. The reversible fluorescence activation, possible in one instance, imposes intermittence on the detected emission and offers the opportunity to resolve closely-spaced nanocarriers in time to reconstruct images with subdiffraction resolution. The irreversible fluorescence activation, possible in the other, maintains emission on after the activation event and permits the monitoring of the diffusion of the activated nanocarriers in real time with the sequential acquisition of images. Thus, these operating principles to solubilize and operate photoswitchable fluorophores in aqueous environments with the aid of supramolecular nanocarriers can lead to valuable protocols to image specimens with subdiffraction resolution and to monitor dynamic events noninvasively.}, booktitle={Colloidal Nanoparticles for Biomedical Applications XI}, publisher={SPIE-Intl Soc Optical Eng}, author={Zhang, Yang and Tang, Sicheng and Thapaliya, Ek Raj and Raymo, Franҫisco M.}, editor={Parak, Wolfgang J. and Osinski, Marek and Liang, Xing-JieEditors}, year={2016}, month={Apr} }
 @article{ragab_thapaliya_zhang_tang_mcmahan_syed_captain_raymo_2016, title={Synthesis in living cells with the assistance of supramolecular nanocarriers}, volume={6}, DOI={10.1039/c6ra04335f}, abstractNote={Supramolecular nanocarriers transport complementary reactants inside living cells in consecutive internalization steps to allow their reaction exclusively in the intracellular space with the formation of a fluorescent product.}, number={39}, journal={RSC Adv.}, publisher={Royal Society of Chemistry (RSC)}, author={Ragab, Sherif Shaban and Thapaliya, Ek Raj and Zhang, Yang and Tang, Sicheng and McMahan, Jeffrey Blye and Syed, Sheyum and Captain, Burjor and Raymo, Françisco M.}, year={2016}, pages={32441–32445} }
 @article{zhang_garcia-amorós_captain_raymo_2016, title={A fluorescent and halochromic indolizine switch}, volume={4}, DOI={10.1039/c5tc03331d}, abstractNote={Proton transfer from a photoacid generator to a halochromic molecular switch within a polymer film permits the imprinting of fluorescent patterns under the influence of optical stimulations.}, number={14}, journal={J. Mater. Chem. C}, publisher={Royal Society of Chemistry (RSC)}, author={Zhang, Yang and Garcia-Amorós, Jaume and Captain, Burjor and Raymo, Françisco M.}, year={2016}, pages={2744–2747} }
 @article{thapaliya_fowley_callan_tang_zhang_callan_raymo_2015, title={Energy-Transfer Schemes To Probe Fluorescent Nanocarriers and Their Emissive Cargo}, volume={31}, DOI={10.1021/acs.langmuir.5b01949}, abstractNote={A strategy to probe supramolecular nanocarriers and their cargo in the intracellular space was developed on the basis of fluorescence measurements and energy transfer. It relies on the covalent attachment of an energy donor, or acceptor, to the macromolecular backbone of amphiphilic polymers and the noncovalent encapsulation of a complementary acceptor, or donor, in the resulting micelles. In aqueous environments, these macromolecules self-assemble into nanostructured constructs and bring the complementary chromophores in close proximity to enable efficient energy transfer. These supramolecular assemblies travel from the extracellular to the intracellular space and retain their integrity in the process. Indeed, donors and acceptors remain close to each other after internalization, and excitation of the former chromophores translates into significant intracellular emission from the latter. Furthermore, these supramolecular assemblies exchange their components with fast kinetics in aqueous dispersions because of the reversible character of the noncovalent contacts holding them together. As a result, micelles incorporating exclusively the donors and nanocarriers containing only the acceptors scramble their chromophoric building blocks, upon mixing, to allow the transfer of energy. These dynamic processes can be reproduced in the intracellular environment with the sequential incubation of cells with the two sets of complementary nanostructured assemblies. Thus, these operating principles and choice of supramolecular synthons are particularly valuable to monitor self-assembling nanocarriers and their cargo inside living cells and can facilitate the elucidation of the behavior of these promising delivery vehicles in a diversity of biological specimens.}, number={35}, journal={Langmuir}, publisher={American Chemical Society (ACS)}, author={Thapaliya, Ek Raj and Fowley, Colin and Callan, Bridgeen and Tang, Sicheng and Zhang, Yang and Callan, John F. and Raymo, Françisco M.}, year={2015}, month={Sep}, pages={9557–9565} }
 @article{garcia-amorós_swaminathan_zhang_nonell_raymo_2015, title={Optical writing and reading with a photoactivatable carbazole}, volume={17}, DOI={10.1039/c5cp01336d}, abstractNote={Fluorescence photoactivation of a carbazole switch under plasmonic assistance permits the optical writing and reading of microstructured patterns.}, number={17}, journal={Phys. Chem. Chem. Phys.}, publisher={Royal Society of Chemistry (RSC)}, author={Garcia-Amorós, Jaume and Swaminathan, Subramani and Zhang, Yang and Nonell, Santi and Raymo, Françisco M.}, year={2015}, pages={11140–11143} }
 @article{zhang_swaminathan_tang_garcia-amorós_boulina_captain_baker_raymo_2015, title={Photoactivatable BODIPYs Designed To Monitor the Dynamics of Supramolecular Nanocarriers}, volume={137}, DOI={10.1021/ja5125308}, abstractNote={Self-assembling nanoparticles of amphiphilic polymers can transport hydrophobic molecules across hydrophilic media and, as a result, can be valuable delivery vehicles for a diversity of biomedical applications. Strategies to monitor their dynamics noninvasively and in real time are, therefore, essential to investigate their translocation within soft matrices and, possibly, rationalize the mechanisms responsible for their diffusion in biological media. In this context, we designed molecular guests with photoactivatable fluorescence for these supramolecular hosts and demonstrated that the activation of the fluorescent cargo, under optical control, permits the tracking of the nanocarrier translocation across hydrogel matrices with the sequential acquisition of fluorescence images. In addition, the mild illumination conditions sufficient to implement these operating principles permit fluorescence activation within developing Drosophila melanogaster embryos and enable the monitoring of the loaded nanocarriers for long periods of time with no cytotoxic effects and no noticeable influence on embryogenesis. These photoresponsive compounds combine a borondipyrromethene (BODIPY) chromophore and a photocleavable oxazine within their covalent skeleton. Under illumination at an appropriate activation wavelength, the oxazine ring cleaves irreversibly to bring the adjacent BODIPY fragment in conjugation with an indole heterocycle. This structural transformation shifts bathochromically the BODIPY absorption and permits the selective excitation of the photochemical product with concomitant fluorescence. In fact, these operating principles allow the photoactivation of BODIPY fluorescence with large brightness and infinite contrast. Thus, our innovative structural design translates into activatable fluorophores with excellent photochemical and photophysical properties as well as provides access to a general mechanism for the real-time tracking of supramolecular nanocarriers in hydrophilic matrices.}, number={14}, journal={J. Am. Chem. Soc.}, publisher={American Chemical Society (ACS)}, author={Zhang, Yang and Swaminathan, Subramani and Tang, Sicheng and Garcia-Amorós, Jaume and Boulina, Marcia and Captain, Burjor and Baker, James D. and Raymo, Françisco M.}, year={2015}, month={Apr}, pages={4709–4719} }
 @article{garcia-amorós_tang_zhang_thapaliya_raymo_2016, title={Self-Assembling Nanoparticles of Amphiphilic Polymers for In Vitro and In Vivo FRET Imaging}, DOI={10.1007/978-3-319-22942-3_2}, abstractNote={Self-assembling nanoparticles of amphiphilic polymers are viable delivery vehicles for transporting hydrophobic molecules across hydrophilic media. Noncovalent contacts between the hydrophobic domains of their macromolecular components are responsible for their formation and for providing a nonpolar environment for the encapsulated guests. However, such interactions are reversible and, as a result, these supramolecular hosts can dissociate into their constituents amphiphiles to release the encapsulated cargo. Operating principles to probe the integrity of the nanocarriers and the dynamic exchange of their components are, therefore, essential to monitor the fate of these supramolecular assemblies in biological media. The co-encapsulation of complementary chromophores within their nonpolar interior offers the opportunity to assess their stability on the basis of energy transfer and fluorescence measurements. Indeed, the exchange of excitation energy between the entrapped chromophores can only occur if the nanoparticles retain their integrity to maintain donors and acceptors in close proximity. In fact, energy-transfer schemes are becoming invaluable protocols to elucidate the transport properties of these fascinating supramolecular constructs in a diversity of biological preparations and can facilitate the identification of strategies to deliver contrast agents and/or drugs to target locations in living organisms for potential diagnostic and/or therapeutic applications.}, journal={Light-Responsive Nanostructured Systems for Applications in Nanomedicine}, publisher={Springer Science \mathplus Business Media}, author={Garcia-Amorós, Jaume and Tang, Sicheng and Zhang, Yang and Thapaliya, Ek Raj and Raymo, Françisco M.}, year={2016}, pages={29–59} }
 @article{cui_zhang_zhou_yu_2013, title={Two-Photon Absorption Properties of <i>s</i>-Triazine Derivatives With Near Octupolar Symmetry}, volume={652-654}, DOI={10.4028/www.scientific.net/amr.652-654.542}, abstractNote={Two new three-branched s-triazine derivatives with near-octupolar symmetry have been synthesized. In each of them, two branches have donor groups attaching to their ends, while the other one has none. Their photophysical characteristics, such as UV–vis spectra, fluorescence spectra and two-photon excited fluorescence spectra, are compared with their octuplar analogs in which all the three branches are ended by donor group. The little structural difference results in great decline of single-photon absorptivity and single-photon excited fluorescence, as well as the two-photon excitated fluorescence.}, journal={AMR}, publisher={Trans Tech Publications}, author={Cui, Yue Zhi and Zhang, Yang and Zhou, Long Long and Yu, Fang}, year={2013}, month={Jan}, pages={542–545} }
 @article{peng_cui_zhang_zhang_guan_2011, title={Optical Properties of Chromophores with Different Six-Membered N-Heterocyclic Aromatic Ring}, volume={236-238}, DOI={10.4028/www.scientific.net/amr.236-238.1598}, abstractNote={A series of intramolecular charge transfer (ICT) molecules containing six-membered N-heterocyclic electron acceptors, as well as their phenyl analogue, have been synthesized. The pyrazine derivatives exhibit the largest quantum yield, stokes shift and the longest emission maxima wavelength due to the appropriate electronegativity of pyrazine, while the triazine derivative has much lower quantum yield due to its too strong ICT. The methyl groups at N-heterocyclic rings also have influence on the optical properties by disturbing the molecular dipole moment.}, journal={AMR}, publisher={Trans Tech Publications}, author={Peng, Hai Tao and Cui, Yue Zhi and Zhang, Tao and Zhang, Yang and Guan, Hai Xing}, year={2011}, month={May}, pages={1598–1602} }