Molecular Engineering and Design of Semiconducting Polymer Dots with NarrowBand, Near-Infrared Emission for in Vivo Biological Imaging Chi-Shiang Ke,† Chia-Chia Fang,† Jia-Ying Yan,‡ Po-Jung Tseng,† Joseph R. Pyle,§ Chuan-Pin Chen,† Shu-Yi Lin,‡ Jixin Chen,§ Xuanjun Zhang,*,⊥ and Yang-Hsiang Chan*,† †
Department of Chemistry, National Sun Yat-sen University, 70 Lien Hai Road, Kaohsiung, Taiwan 80424 Center for Nanomedicine Research, National Health Research Institutes, 35 Keyan Road, Zhunan, Taiwan 35053 § Department of Chemistry & Biochemistry, Ohio University, Athens, Ohio 45701, United States ⊥ Faculty of Health Sciences, University of Macau, Macau SAR, China ‡
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ABSTRACT: This article describes the design and synthesis of donor−bridge−acceptor-based semiconducting polymer dots (Pdots) that exhibit narrow-band emissions, ultrahigh brightness, and large Stokes shifts in the near-infrared (NIR) region. We systematically investigated the effect of π-bridges on the fluorescence quantum yields of the donor−bridge−acceptor-based Pdots. The Pdots could be excited by a 488 or 532 nm laser and have a high fluorescence quantum yield of 33% with a Stokes shift of more than 200 nm. The emission full width at half-maximum of the Pdots can be as narrow as 29 nm, about 2.5 times narrower than that of inorganic quantum dots at the same emission wavelength region. The average per-particle brightness of the Pdots is at least 3 times larger than that of the commercially available quantum dots. The excellent biocompatibility of these Pdots was demonstrated in vivo, and their specific cellular labeling capability was also approved by different cell lines. By taking advantage of the durable brightness and remarkable stability of these NIR fluorescent Pdots, we performed in vivo microangiography imaging on living zebrafish embryos and long-term tumor monitoring on mice. We anticipate these donor−bridge−acceptor-based NIR-fluorescent Pdots with narrow-band emissions to find broad use in a variety of multiplexed biological applications. KEYWORDS: semiconducting polymer dots, narrow fluorescence, near-infrared emission, π-bridge, bioimaging ptical imaging techniques with fluorescence microscopy possess numerous advantages including high signal-to-noise ratio, excellent spatial and temporal resolution, and noninvasive nature, making them extremely useful in the investigation of biological processes in live organisms. Recently, the exciting advances in fluorescence image-guided surgery1,2 and its first in-human proof-of-concept staging and ovarian debulking surgery 3,4 have greatly emphasized the significance of selecting a bright, photostable, biocompatible, and tumor-targetable fluorescent probe. Over the past few years, the use of visible to invisible near-infrared (NIR) fluorescent light in intraoperative imaging has had a considerable role to fill the gap between preoperative staging and intraoperative planning of resection.5,6 Whereas visible light can travel only several micrometers in tissues, NIR light (700− 900 nm) can penetrate several millimeters (up to centimeters)
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through blood or tissues and cause minimal tissue damage to precisely guide surgeons in real time. Within the NIR window, most biological species such as oxyhemoglobin, deoxyhemoglobin, water, melanon, and lipid absorb minimal light and exhibit low scattering.7,8 Moreover, tissues have almost no autofluorescence inside the NIR spectrum, and thus a high signal-to-noise contrast can be achieved using NIR-responsive fluorescent probes. To date, unfortunately, most FDA-approved clinical agents4 (e.g., indocyanine green, 5-aminolevulinic acid, and methylene blue) suffer from irreversible photobleaching and issues of low brightness and small Stokes shifts. As a result, the development of well-designed NIR fluorophores with high Received: January 10, 2017 Accepted: February 21, 2017 Published: February 21, 2017 3166
DOI: 10.1021/acsnano.7b00215 ACS Nano 2017, 11, 3166−3177
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Design and Synthesis of π-Bridge Spacers. There are three important functions needed to be considered for the design of the π-bridges: (i) to gain the absorption in the visible regions (400−600 nm); (ii) to increase the steric effect of the semiconducting polymers and thereby alleviate the phenomenon of aggregation-caused quenching in a nanoparticle form; (iii) to ameliorate energy transfer from the donor segment to the acceptor moiety. Here we designed four different types of π-bridges, TBT, TAZ, TBTOC6, and TC6FQ, as shown in Figure 1. In our previous studies, we have proven that the
brightness, good photostability, and large Stokes shifts is a profound need in modern bioimaging and clinical diagnosis. In the past decade, nanoparticle-based fluorescent probes such as dye-doped silica/polymeric nanoparticles9−13 and quantum dots14−16 have appeared to be promising candidates due to their remarkably higher brightness and photostability as compared to conventional organic dyes. The potential leaking of the toxic elements (e.g., Cd) from the quantum dots17−19 and the leaching of the embedded materials from the polymeric/silica cores,20,21 however, could seriously hinder their biological applications and clinical implementation. Very recently, semiconducting polymer nanoparticles (Pdots) with compact sizes (
