Image guided photodynamic therapy (PDT) combines fluorescence tracing and phototherapy, which can achieve a more accurate and effective treatment effect. However, traditional photosensitizers are limited by the aggregation-caused fluorescence quenching (ACQ) effect and low reactive oxygen species (ROS) generation in a hypoxic environment, resulting in poor imaging and treatment effect. Herein, we report a tricyano-methylene-pyridine (TCM)-based Type I aggregation-induced emission (AIE) photosensitizer (TCM-MBP), the strong electron acceptance (D-A) effect extends the wavelength to near-infrared (NIR) region to reduce the autofluorescence interference, and oxygen atoms provide lone pair electrons to enhance the inter system crossing (ISC) rate, thereby promoting the generation of more triplet states to produce ROS. The AIE photosensitizer TCM-MBP exhibited low oxygen dependence, NIR emission, and higher ROS production compared to commercially available Ce 6 and RB. After encapsulation with DSPE-PEG2000, TCM-MBP nanoparticles (TCM-MBP NPs) could penetrate to visualize cells and efficiently kill cancer cells upon light irradiation. This study provides an oxygen-independent AIE photosensitizer, which has great potential to replace the commercial ACQ photosensitizers.
The twisted donor-acceptor (D-A) organic formwork with a large dihedral angle (θDA ) is usually adopted to narrow the singlet-triplet energy gap for obtaining excellent thermally activated delayed fluorescence (TADF) emitters. However, the dependence of overall TADF properties on θDA has not been systematically investigated to this day. Taking new designed CzBP, CzBP-1M and CzBP-2M via introducing methyl as investigated models, it is found that (i) with increasing θDA , the charge transfer component in S1 is larger than that in T1 in varying degrees, leading to non-monotonic spin-orbit couplings; (ii) the electron-vibration couplings between S1 and T1 states become the largest when θDA approaching 80°, facilitating phonon-driven up-conversion; (iii) the overall TADF rate reaches a peak at θDA ≈80°. By this, the TADF on/off switching is realized via methyl moiety for regulating θDA from theoretical prediction to experimental confirmation. Importantly, the θDA near 80° would be a good descriptor for screening excellent D-A type TADF emitters.
Abstract Pure organic materials with ultralong room‐temperature phosphorescence (RTP) are attractive alternatives to inorganic phosphors. However, they generally show inefficient intersystem crossing (ISC) owing to weak spin–orbit coupling (SOC). A design principle based on the realization of small energy gap between the lowest singlet and triplet states (ΔE ST ) and pure ππ* configuration of the lowest triplet state (T 1 ) via structural isomerism was used to obtain efficient and ultralong RTP materials. The meta isomer of carbazole‐substituted methyl benzoate exhibits an ultralong lifetime of 795.0 ms with a quantum yield of 2.1 %. Study of the structure–property relationship shows that the varied steric and conjugation effects imposed by ester substituent at different positions are responsible for the small ΔE ST and pure ππ* configuration of T 1 .
Open AccessCCS ChemistryRESEARCH ARTICLE1 May 2022Spatiotemporal Visualization of Cell Membrane with Amphiphilic Aggregation-Induced Emission-Active Sensor Youheng Zhang, Qi Wang, Zhirong Zhu, Weijun Zhao, Chenxu Yan, Zhenxing Liu, Ming Liu, Xiaolei Zhao, He Tian and Wei-Hong Zhu Youheng Zhang Key Laboratory for Advanced Materials and Joint International Research Laboratory of Precision Chemistry and Molecular Engineering, Feringa Nobel Prize Scientist Joint Research Center, Shanghai Key Laboratory of Functional Materials Chemistry, Institute of Fine Chemicals, Frontiers Science Center for Materiobiology and Dynamic Chemistry, School of Chemistry and Molecular Engineering, East China University of Science & Technology, Shanghai 200237 , Qi Wang *Corresponding authors: E-mail Address: [email protected] E-mail Address: [email protected] Key Laboratory for Advanced Materials and Joint International Research Laboratory of Precision Chemistry and Molecular Engineering, Feringa Nobel Prize Scientist Joint Research Center, Shanghai Key Laboratory of Functional Materials Chemistry, Institute of Fine Chemicals, Frontiers Science Center for Materiobiology and Dynamic Chemistry, School of Chemistry and Molecular Engineering, East China University of Science & Technology, Shanghai 200237 , Zhirong Zhu Key Laboratory for Advanced Materials and Joint International Research Laboratory of Precision Chemistry and Molecular Engineering, Feringa Nobel Prize Scientist Joint Research Center, Shanghai Key Laboratory of Functional Materials Chemistry, Institute of Fine Chemicals, Frontiers Science Center for Materiobiology and Dynamic Chemistry, School of Chemistry and Molecular Engineering, East China University of Science & Technology, Shanghai 200237 , Weijun Zhao Key Laboratory for Advanced Materials and Joint International Research Laboratory of Precision Chemistry and Molecular Engineering, Feringa Nobel Prize Scientist Joint Research Center, Shanghai Key Laboratory of Functional Materials Chemistry, Institute of Fine Chemicals, Frontiers Science Center for Materiobiology and Dynamic Chemistry, School of Chemistry and Molecular Engineering, East China University of Science & Technology, Shanghai 200237 , Chenxu Yan Key Laboratory for Advanced Materials and Joint International Research Laboratory of Precision Chemistry and Molecular Engineering, Feringa Nobel Prize Scientist Joint Research Center, Shanghai Key Laboratory of Functional Materials Chemistry, Institute of Fine Chemicals, Frontiers Science Center for Materiobiology and Dynamic Chemistry, School of Chemistry and Molecular Engineering, East China University of Science & Technology, Shanghai 200237 , Zhenxing Liu Key Laboratory for Advanced Materials and Joint International Research Laboratory of Precision Chemistry and Molecular Engineering, Feringa Nobel Prize Scientist Joint Research Center, Shanghai Key Laboratory of Functional Materials Chemistry, Institute of Fine Chemicals, Frontiers Science Center for Materiobiology and Dynamic Chemistry, School of Chemistry and Molecular Engineering, East China University of Science & Technology, Shanghai 200237 , Ming Liu Key Laboratory for Advanced Materials and Joint International Research Laboratory of Precision Chemistry and Molecular Engineering, Feringa Nobel Prize Scientist Joint Research Center, Shanghai Key Laboratory of Functional Materials Chemistry, Institute of Fine Chemicals, Frontiers Science Center for Materiobiology and Dynamic Chemistry, School of Chemistry and Molecular Engineering, East China University of Science & Technology, Shanghai 200237 , Xiaolei Zhao Key Laboratory for Advanced Materials and Joint International Research Laboratory of Precision Chemistry and Molecular Engineering, Feringa Nobel Prize Scientist Joint Research Center, Shanghai Key Laboratory of Functional Materials Chemistry, Institute of Fine Chemicals, Frontiers Science Center for Materiobiology and Dynamic Chemistry, School of Chemistry and Molecular Engineering, East China University of Science & Technology, Shanghai 200237 , He Tian Key Laboratory for Advanced Materials and Joint International Research Laboratory of Precision Chemistry and Molecular Engineering, Feringa Nobel Prize Scientist Joint Research Center, Shanghai Key Laboratory of Functional Materials Chemistry, Institute of Fine Chemicals, Frontiers Science Center for Materiobiology and Dynamic Chemistry, School of Chemistry and Molecular Engineering, East China University of Science & Technology, Shanghai 200237 and Wei-Hong Zhu *Corresponding authors: E-mail Address: [email protected] E-mail Address: [email protected] Key Laboratory for Advanced Materials and Joint International Research Laboratory of Precision Chemistry and Molecular Engineering, Feringa Nobel Prize Scientist Joint Research Center, Shanghai Key Laboratory of Functional Materials Chemistry, Institute of Fine Chemicals, Frontiers Science Center for Materiobiology and Dynamic Chemistry, School of Chemistry and Molecular Engineering, East China University of Science & Technology, Shanghai 200237 https://doi.org/10.31635/ccschem.021.202100967 SectionsSupplemental MaterialAboutAbstractPDF ToolsAdd to favoritesDownload CitationsTrack Citations ShareFacebookTwitterLinked InEmail High-fidelity spatiotemporal monitoring of the cell membrane is critically important. However, commercial fluorescence probes are stalked by the aggregation-caused quenching (ACQ) effect, and the reported aggregation-induced emission (AIE)-active probes are always limited by nonspecific aggregations in the biological environment. Herein, we report the rational molecular design of a state-of-the-art amphiphilic AIE luminogen (AIEgen), membrane tracker QMC12, using a core quinoline-malononitrile (QM) structure to suppress the ACQ effect, incorporate a positively charged pyridinium to regulate dispersity and strengthen the binding affinity to the negatively charged cell membrane, and extend the alky chain to improve the anchoring ability to the cell membrane. The membrane tracker QMC12, which disperses well in both hydrophilic and lipophilic environments, not only achieves minimal background interference and high signal-to-noise (S/N) ratio in the "ultrafast" visualization of the cell membrane, but also endows a "wash-free" characteristic. Furthermore, it realizes a spatial three dimensional (3D) view in a multicellular spheroid model and morphology changes over time. Moreover, QMC12 avoids false staining and signal loss and unprecedentedly achieves the direct observation of the cell membrane's microstructure, which could elucidate spatiotemporal 3D model studies of the intercellular information exchange. Download figure Download PowerPoint Introduction The cell membrane, mainly composed of phospholipid bilayers, has a tremendous role in accurately coordinating various cellular behaviors.1–6 It is critically important to monitor the spatiotemporal changes of the cell membrane for early medical diagnosis and basic biological research.7–9 However, commercial fluorescence probes such as DiO or Dil are not ideal for cell membrane imaging because of their inherent aggregation-caused quenching (ACQ) effect, which often causes false signals and inevitable noises from an "always-on" pattern.10–13 Moreover, these commercial probes show poor solubility in both aqueous and lipid solutions, which complicates the staining procedures and limits further application at the multicellular level. In addition, their low positive charge density and weak hydrophobic interaction sometimes result in poor targetability and weak anchoring ability, thus providing inaccurate information feedback such as signal loss and unsustainable imaging (Figure 1a). Figure 1 | Rational design of the unique amphiphilic AIE-active probe for high-fidelity spatiotemporal mapping of the cell membrane. (a) Commercial probe DiO and Dil based on "always-on" pattern. (b) The rational design of the amphiphilic AIE-active probe to overcome the inherent deficiencies of commercial probes. The dimethylamino benzene group was used to extend wavelength, the pyridinium salt group for regulating solubility and targetability, and the long alkyl chain for promoting the anchoring property to cell membrane. (C) Left: schematic illustration of amphiphilic AIE-active QMC12 labelling cell membrane, thereby achieving "off–on" fluorescence upon enrichment in the cell membrane. Right: the interaction between the amphiphilic AIE-active probe QMC12 and a phospholipid molecule. Download figure Download PowerPoint Although design of fluorescence probes employing the concept of aggregation-induced emission (AIE) is highly desirable to overcome the "always on" pattern,14–24 most reported AIE luminogen (AIEgen)based probes still suffer from unexpected aggregations owing to their poor solubility in either aqueous or lipid environments, thereby providing inaccurate fluorescence signal. To resolve these issues, the water solubility of probes based on AIEgens must be improved to enable good dispersity in aqueous environment, decrease the background signal, simplify the operation procedure, and improve the multicellular level staining. Meanwhile, enhancement of the lipid solubility is expected to avoid false signals created by unexpected aggregation in lipid organelles. Furthermore, a positively charged group could be introduced, and the hydrophilicity and hydrophobicity can be adjusted to improve the affinity with the amphiphilic phospholipid bilayer to achieve targetability and anchoring ability to the cell membrane. In this work, we describe a rational design strategy to construct a novel amphiphilic AIE-active probe with a strong targeting ability and anchoring property for high-fidelity spatiotemporal imaging of the cell membrane. This amphiphilic AIEgen relies on the quinoline-malononitrile (QM) building block to overcome the ACQ effect, introduce the positively charged pyridinium salt to regulate the aggregation behavior in hydro- and lipophilic environments and generate strong binding affinity, and extend the alkyl chain to adjust the hydrophilicity and hydrophobicity (Figure 1b). With this strategy, the elaborated membrane tracker QMC12 posesses the following extraordinary features: (1) achieves high signal-to-noise (S/N) ratio with amphiphilic AIEgens through overcoming the ACQ effect and eliminating the undesired aggregations in hydro- and lipophilic environments; (ii) avoids false staining and signal loss with assistance of the superior targeting aggregation and excellent anchoring ability; (3) realizes ultrafast "wash-free" imaging because of superior water solubility with facilitated staining procedure; (4) spatiotemporally stains multicellular models because of the good dispersity and beneficial diffusion between cell membranes; and (5) maps morphology change over time with strong anchoring ability by tuning hydrophilicity and hydrophobicity to phospholipid bilayers (Figure 1c). In summary, the amphiphilic QMC12 has for the first time achieved 2D and 3D spatiotemporal visualization of the cell membrane without signal loss or false staining, even clearly observing the neurons dendrites' microstructure, thus providing a promising alternative to commercial probes such as DiO or Dil for cell membrane imaging. Experimental Methods Materials and general methods All solvents and chemicals, unless specifically stated, were purchased commercially in analytical grade and used without further purification. 1H and 13C NMR spectra in deuterated solvent were obtained with a Bruker AvanceIII 400 MHz NMR spectrometer (Billerica, MA) using tetramethylsilane (TMS) as an internal standard. High-resolution mass spectrometry (HRMS) spectra were measured with a Waters LCT Premier XE spectrometer (Billerica, MA). Synthesis of compounds First, Py-QM was prepared by coupling the pyridine group with Br-QM through Suzuki reaction. Then, QM-PN and Py-QM-PN were synthesized by Knoevenagel condensation of QM and Py-QM with 4-dimethylaminobenzaldehyde. Finally, Py-QM-PN was treated with iodoethane and 1-iodododecane to obtain QMC2 and QMC12 after ion exchange. The detailed synthesis routes of all compounds were shown in Supporting Information Scheme S1. Cell lines The adenocarcinoma human alveolar basal epithelial cells (A549), human pancreatic cancer cell (PANC-1), adrenal pheochromocytoma cell (PC12), and human epithelioid cervical carcinoma cell (HeLa) were purchased from the Institute of Cell Biology (Shanghai, China). Cells were all propagated in T-75 flasks cultured at 37 °C under a humidified 5% CO2 atmosphere in Dulbecco's modified Eagle's medium (DMEM) (GIBCO/Invitrogen, Camarillo, CA), which was supplemented with 10% fetal bovine serum (FBS; Biological Industry, Kibbutz Beit Haemek, Israel) and 1% penicillin–streptomycin (10,000 U mL−1 penicillin and 10 mg mL−1 streptomycin; Solarbio Life Science, Beijing, China). In vitro cytotoxicity assay The cell cytotoxicity of QMC2 and QMC12 in HeLa cells was evaluated by a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Briefly, cells were seeded into 96-well plates at a density of 1 × 104 cells/well and cultured at 37 °C under a humidified 5% CO2 atmosphere for 12 h. Then, the cells were exposed to various concentrations (1, 2.5, 5.0, 7.5, and 10 μM) of QMC2 and QMC12 or 100 μL culture medium as a negative control group. After incubation at 37 °C under a humidified 5% CO2 atmosphere for 24 h, MTT solution (5 mg/mL, 10 μL) was added to the media and incubated for another 4 h, and the absorbance at 490 nm was measured with a Multimode Plate Reader (BioTek, Burlington, VT). The relative cell viability (%) was calculated by the following formula: Cell viability (%) = mean absorbance value of the treatment group-blank/mean absorbance value of the control blank × 100. Cell imaging HeLa cells were seeded onto glass-bottom Petri dishes in culture medium (1.0 mL) and allowed to adhere for 12 h before imaging. Probe QM-PN, QMC2, and QMC12 at a final concentration of 5 × 10−6 M [containing 0.1% dimethyl sulfoxide (DMSO)] were added into culture medium and incubated for different time at 37 °C under a humidified 5% CO2 atmosphere. Cell imaging was captured by using a confocal laser scanning microscope (CLSM, Leica TCS SP8) with a 63× oil immersion objective lens. The fluorescence signals of cells incubated with probes were collected at 600–750 nm under excitation wavelength at 561 nm. Results and Discussion Incorporating pyridinium salt and long alkyl chain for amphiphilic AIE-active sensor To avoid the premature activated fluorescence caused by nonspecific aggregations in reported AIE sensors,25,26 we rationally designed and constructed an amphiphilic AIE-active probe step-by-step.27,28 First, the newly developed AIE building block, QM, was employed as the core structure to overcome the ACQ effect of commercial probes ( Supporting Information Figure S1), and the π-conjugated backbone dimethylamino benzene was utilized to extend the emission wavelength to provide QM-PN.29–33 Then, the electron-withdrawing pyridine group was introduced to further extend emission wavelength, affording the intermediate Py-QM-PN. As we expected, a significant fluorescence red shift of 111 nm was observed from QM to Py-QM-PN at solid state ( Supporting Information Figure S2). Sequentially, the ethyl group was covalently connected to the pyridine group, thus achieving positively charged QMC2, which was supposed to control the specific solubility in both lipophilic and hydrophilic systems with an initial "fluorescence-off" state and realize high targetability to the negativelycharged cell membrane through electron interaction.34–36 Finally, we extended the alkyl chain to tune the hydrophilicity and hydrophobicity, yielding membrane tracker QMC12, in which the anchoring ability could be strengthened by better hydrophobic interaction between the probe molecule and phospholipid bilayer of the cell membrane (Figure 1c). The molecular structures of all the designed compounds were confirmed by 1H NMR, 13C NMR, and HRMS. Amphiphilicity significantly minimizes undesirable fluorescence signal Previously developed AIEgen-based probes disperse well in either aqueous or lipid environments and thus are unable to achieve real targeting of the cell membrane; therefore, an amphiphilic AIE probe is expected to change this undesirable situation. To investigate the AIE and amphiphilicity properties of QM-PN and Py-QM-PN, their fluorescence spectra were first investigated in mixed water/tetrahydrofuran (THF) solvents with different fractions of water (fw). As shown in Figures 2a and 2b, the fluorescence intensity was enhanced when the water fraction gradually increased for both QM-PN (fw from 0% to 70%) and Py-QM-PN (fw from 0% to 60%), suggestive of their significant AIE property. The fluorescence intensity slightly decreased when the water fraction increased to 99%, which we attributed to the formation of nanoaggregates with a looser packing mode (Figure 2c). Both dynamic light scattering (DLS) (Figure 2d) and transmission electron microscopy (TEM) ( Supporting Information Figure S3) further supported that nanoaggregations formed in the aqueous environment (99% water). Obviously, QM-PN and Py-QM-PN inherit the excellent AIE features of the QM core, but they are largely hydrophobic and can form nonspecific aggregations in the aqueous system, thus still suffering from the "always-on" pattern for cell membrane imaging. Figure 2 | QMC12 exhibited "off–on" characteristic with amphiphilic behavior. (a–c) Fluorescence emission spectra and I/I0 plots of QM-PN (λex = 431 nm) and Py-QM-PN (λex = 442 nm) in a mixture of THF/water with different fw. I0 is the fluorescence intensity of QM-PN and Py-QM-PN in 0% water. (d) Size distribution of QM-PN and Py-QM-PN in a mixture of DMSO/Water (v/v = 1/99) obtained from DLS. Emission spectra of (e) QMC2 (10 μM) and (i) QMC12 (10 μM) in THF/water (λex = 447 nm). Emission spectra and I/I0 plots of (f and g) QMC2 and (j and k) QMC12 in Gly/water system with different Gly fractions (fg), λex = 454 nm. I0 is the fluorescence intensity of QMC2 and QMC12 in 99% water. Inset: fluorescence photographs of QMC2 and QMC12 in fg = 0 and fg = 99 taken under 365 nm UV irradiation. Size distribution of (h) QMC2 and (l) QMC12 (10 μM) in different solvents obtained from DLS. (m) HOMO and LUMO of QM-PN and QMC12 by DFT calculations. Download figure Download PowerPoint Compared with hydrophobic AIEgens (QM-PN and Py-QM-PN), QMC2 and QMC12, modified by a hydrophilic pyridinium salt unit, exhibited desirable amphiphilicity. Both showed extremely weak emission in all THF/water (Figures 2e and 2i), DMSO/water ( Supporting Information Figure S4), ethanol/water ( Supporting Information Figure S5) at any fraction of water, phosphate-buffered saline, and 10% FBS solution ( Supporting Information Figure S6). Those initial "fluorescence-off" states may result from their good dispersity in both aqueous and organic solvents, wherein QMC2 and QMC12 have free intramolecular motions and thus cause the fluorescence quenching according to the classic AIE mechanism. Moreover, DLS (Figures 2h and 2l) and TEM ( Supporting Information Figure S3) results show that QMC2 and QMC12 had extremely small sizes in both hydrophilic and lipophilic environments, proving their amphiphilic character. The AIE behaviors of QMC2 (Figures 2f and 2g) and QMC12 (Figures 2j and 2k) were further investigated in a glycerin (Gly)/water system through simulating the restriction of intramolecular motion (RIM) in high-viscosity environments. Both probes are non-emissive in aqueous solutions, but with increasing viscosity (fraction of Gly, fg = 0–99%), the fluorescence intensity gradually increased 31.2 times for QMC2 (Figure 2g) and 35.3 times for QMC12 (Figure 2k) via eliminating the non-radiative channel with the specific RIM mechanism. These results demonstrate that the unique amphiphilic QMC2 and QMC12 inherited the AIE property from the QM core and achieved the "fluorescence off" state in an aqueous system until it encountered the targeted site, which restricts the intramolecular motion, thus achieving high-fidelity imaging with a fluorescence "off–on" response. The wavelength extension from QM to QMC12 was realized by enhancing the electronic donor–acceptor effect, in which the absorption successfully shifted from 414 nm of QM to 455 nm of QMC12, and the emission peak red shifted from 514 nm of QM to 628 nm of QMC12 ( Supporting Information Figure S7).28 According to density functional theory (DFT) calculations the highest occupied molecular orbital (HOMO) electron density of QMC12 is mainly delocalized at the electron-rich N,N′-dimethylamino unit and phenyl group, and the lowest unoccupied molecular orbital (LUMO) is mainly delocalized at the pyridinium salt group (Figure 2m and Supporting Information Figure S8). The smaller HOMO–LUMO energy gap of QMC12 further confirmed the stronger electronic donating–accepting interaction, which is the reason for the longer absorption wavelength. It is highly expected that the amphiphilic probe QMC12 could overcome the ACQ effect from the "always-on" pattern as well as avoid undesirable aggregations in both hydrophilic and hydrophobic environments until encountering the high-viscosity cell membrane to emit fluorescence, thus it achieves "off–on" behavior in mapping the cell membrane. High-fidelity imaging of cell membrane Accurate imaging of the cell membrane has great importance for tracing cell distribution and reflecting the structural integrity.37 Considering the positively charged sensor could accumulate at the negatively charged cell membrane and the long alkyl chain could anchor to the phospholipid bilayers, the QMC12 is expected to provide high-fidelity imaging of cell membrane. Here, all the designed molecules were evaluated by co-staining with commercial membrane tracker DiO in HeLa cells (human epithelioid cervical carcinoma cells). As shown in Figures 3a and 3b, the lipophilic probes QM-PN and Py-QM-PN showed obvious fluorescence signal in the cytoplasm rather than the cell membrane, resulting in poor Pearson's correlation coefficient of 0.04 for QM-PN and 0.12 for Py-QM-PN ( Supporting Information Figure S9). In addition, the intensity profile of the linear region of interest (ROI) across the cells showed that QM-PN and Py-QM-PN do not overlap well with the commercial DiO, further demonstrating the poor co-localization ability (Figures 3a and 3b). These results indicate that the lipophilicity of QM-PN and Py-QM-PN make them more favorably penetrate through the cell membrane and aggregate as AIE nanoparticles to emit fluorescence in the cytoplasm. In contrast, the positively charged QMC2 and QMC12 displayed bright fluorescence signal in the cell membrane rather than the cytoplasm or extracellular matrix, which confirmed the targeting ability of the positively charged sensor towards the negatively charged cell membrane (Figures 3c and 3d). It is noted that the QMC2 with short alkyl chain exhibited a smaller Pearson's correlation coefficient of 0.45 (Figure 3c and Supporting Information Figure S9) than the correlation coefficient of QMC12 (0.88), suggesting a stronger anchoring property with longer alkyl chain (Figure 3d and Supporting Information Figure S9). In addition, several cell lines' cell membranes were stained well using QMC12 probe with wide application range ( Supporting Information Figure S10). These results strongly support the remarkable cell membrane targeting ability of amphiphilic QMC12. Figure 3 | High-fidelity imaging of cell membrane by QMC12. (a–d) HeLa cells were incubated with QM-PN (5 μM), Py-QM-PN (5 μM), QMC2 (5 μM), and QMC12 (5 μM) for 20 min followed by co-staining with DiO (10 μM) for 20 min. (a2–d2) Green channels from DiO (λex = 488 nm, λem = 500–600 nm). (a3–d3) Red channels from QM-PN, Py-QM-PN, QMC2, or QMC12 (λex = 561 nm, λem = 600–750 nm). (a4–d4) Merged images of green, red, and bright field channels. (a5–d5) The intensity profile of the white linear ROI across the cell in a4–d4. All images share the same scale bar of 10 μm. Download figure Download PowerPoint To further confirm that the amphiphilicity of QMC12 can eliminate undesirable aggregations and the proper hydrophilicity and hydrophobicity can ensure strong anchoring ability to phospholipid bilayers, we compared the cell membrane imaging performance of QMC12 with commercial trackers. Due to excessive lipid solubility and poor anchoring ability, both commercial DiO and Dil sensors showed undesired false staining (Figure 4a and Supporting Information Figure S11) and signal loss (Figure 4b). In contrast, amphiphilic QMC12 would not accumulate on bubbles with less electrostatic interaction, thus successfully avoiding a false signal (Figure 4a). Moreover, due to the effective electrostatic interaction and the strong anchoring effect of the long alkyl chain between cell membrane and probe, QMC12 could avoid signal loss after several experimental operations (Figure 4b). These results indicate that QMC12 has realized high-fidelity imaging of cell membrane with high S/N ratio. Figure 4 | High-fidelity 3D imaging of cell membrane. (a) False signal and (b) signal loss of DiO compared with QMC12. All images share the same scale bar of 10 μm. (c) Z-stack images of Hela cells from 0–9 μm after staining with QMC12 (5 μM) for 20 min. All images share the same scale bar of 10 μm. (d) The 3D reconstructed images of Hela cell. (e) Z-stack images of PC12 cells from 0 to 9 μm after staining with QMC12 (5 μM) for 20 min, followed by staining with Hoechst 33342 (20 mM). The overlay channel is merged bright field, blue channel (λex = 405 nm, λem = 420–450 nm), and red channel. All images share the same scale bar of 10 μm. (f) The 3D reconstructed images of PC12 cell. Download figure Download PowerPoint Encouraged by the above cell imaging experiments, more detailed three-dimensional (3D) structural information of the cell membrane was expected to be obtained. Therefore, we recorded a series of confocal images (at different depth) by scanning the HeLa cells (Figures 4c and 4d) and PC12 cells (a common nerve cell line, Figures 4e and 4f) stained with QMC12. As expected, the red fluorescence signal of QMC12 overlapped well with the cell membrane observed in the brightfield images of HeLa cells (Figure 4c) and PC12 cells (Figure 4e), thus showing the 3D morphology as a protective barrier surrounding cells. Surprisingly, the cell membrane was continuous and uninterruptedly stained, even to the extent that the dendrites' microstructure in neuron cell was clearly observed (Figure 4f and Supporting Information Video S1), which characteristic is helpful to observe the interaction between cells, especially the direct information exchange between nerve cells. Taken together, compared to commercial cell membrane trackers, our designed amphiphilic AIE-active probe QMC12 realized high-fidelity 2D and 3D cell membrane imaging with a much higher S/N ratio. Ultra-fast cell membrane staining with "wash-free" behavior The fragility of the cell membrane urgently requires uncomplicated staining procedures. However, the commercial DiO probe (work concentration of 10−3 M) always needs several dissolution and washing procedures because of its poor solubility in both aqueous and organic solvents ( Supporting Information Figure S12).38 The amphiphilic AIEgen QMC12 should disperse well in both aqueous and lipid solutions with an initial "fluorescence-off" property, so as to generate potential "wash-free" behavior, thereby facilitating experimental efficiency and avoiding the signal-loss caused by the postwashing procedure. As shown in Figure 5, obvious fluorescence signal was observed outside the cell membrane after incubating with the DiO for 10 min (Figure 5a), while the extracellular background fluorescence was invisible in both the amphiphilic QMC2 (Figure 5b) and QMC12 (Figure 5c) groups. The normalized fluorescence intensity ratio of the cell membrane to the extracellular matrix is 5-fold for DiO (Figure 5a), 90-fold for the QMC2 group (Figure 5b), and 180-fold for the QMC12 group (Figure 5c). These results demonstrate the post
Influenced by increasing global extreme weather and the uneven spatiotemporal distribution of water resources in monsoon climate areas, the balance of agricultural water resources supply and demand currently faces significant challenges. Conducting research on the spatial allocation trade-offs and synergistic mechanisms of agricultural water resources in monsoon climate areas is extremely important. This study takes the spatial layout of reservoir site selection in water conservancy projects as an example, focusing on Shandong Province as the research area. During the site selection process, the concept of water resource demand is introduced, and the suitability of reservoir siting is integrated. It clarifies ten influencing factors for suitability degree and five influencing factors for demand. A bi-objective optimization model that includes suitability degree and demand degree is established. Utilizing machine learning methods such as the GA_BP neural network model and the GA-bi-objective optimization model to balance and coordinate the supply and demand relationship of agricultural water resources in the monsoon region. The study found that: (1) in the prediction of suitability degree, the influencing factors are most strongly correlated with the regulatory storage capacity (regulatory storage capacity > total storage capacity > regulating storage coefficient); (2) compared with single-objective optimization of suitability degree, the difference between water supply and demand can be reduced by 74.3% after bi-objective optimization; (3) according to the spatial layout optimization analysis, the utilization of water resources in the central and western parts of Shandong Province is not sufficient, and the construction of agricultural reservoirs should be carried out in a targeted manner. This study provides new ideas for promoting the efficient use of water resources in monsoon climate zones and the coordinated development of humans and nature, reflecting the importance of supply and demand balance in the spatial allocation of agricultural water resources, reducing the risk of agricultural production being affected by droughts and floods.
It introduces the motor-driven system of the electrical bicycle.Based on the AVR single chip,through modulating the duty-ratio of PWM wave,the speed of the electrical bicycle can be well controlled.The research is focused on the phenomenon of overcurrent,overload and abnormal operation of the motor during begining.It puts forward the method of overcurrent detecting and protecting.The experimental results show that the method has good ability.
Abstract The development of ultra‐long room‐temperature phosphorescence (UL‐RTP) in processable amorphous organic materials is highly desirable for applications in flexible displays, anti‐counterfeiting, and bio‐imaging. However, achieving efficient UL‐RTP from amorphous materials remains a challenging task, especially with activation by visible light and a bright afterglow. Here we report a general and rational molecular‐design strategy to enable efficient visible‐light‐excited UL‐RTP by multi‐esterification of a rigid large‐plane phosphorescence core. Notably, multi‐esterification minimizes the aggregation‐induced quenching and accomplishes a ′four birds with one stone′ possibility in the generation and radiation process of UL‐RTP: i) shifting the excitation from ultraviolet light to blue‐light through enhancing the transition dipole moment of low‐lying singlet‐states, ii) facilitating the intersystem crossing process through the incorporation of lone‐pair electrons, iii) boosting the decay process of long‐lived triplet excitons resulting from a significantly increased transition dipole moment, and iv) reducing the intrinsic triplet nonradiative decay by substitution of high‐frequency vibrating hydrogen atoms. All these factors synergistically contribute to the most efficient and stable visible‐light‐stimulated UL‐RTP (lifetime up to 2.01 s and efficiency up to 35.4 % upon excitation at 450 nm) in flexible films using multi‐esterified coronene, which allows high‐tech applications in single‐component time‐delayed white light‐emitting diodes and information technology based on flashlight‐activated afterglow encryption.
Stress is critically important for the manufacturing industry. The visualization of stress/strain distributions and fatigue crack propagation on metal specimens with a pure organic fluorescent material is achieved by Zhe Zhang, Ben Zhong Tang, and co-workers in article number 1803924. Such a method enjoys the merits of a simple setup and real-time, full-field, on-site, and direct visualization. The invisible information of the mechanical response of the metal specimens are transformed to visible fluorescent signals.
The development of smart-responsive materials, in particular those with non-invasive, rapid responsive phosphorescence, is highly desirable but has rarely been described. Herein, we designed and prepared a series of molecular rotors containing a triazine core and three bromobiphenyl units: o-Br-TRZ, m-Br-TRZ, and p-Br-TRZ. The bromine and triazine moieties serve as room temperature phosphorescence-active units, and the bromobiphenyl units serve as rotors to drive intramolecular rotation. When irradiated with strong ultraviolet photoirradiation, intramolecular rotations of o-Br-TRZ, m-Br-TRZ, and p-Br-TRZ increase, successively resulting in a photothermal effect via molecular motions. Impressively, the photothermal temperature attained by p-Br-TRZ is as high as 102 °C, and synchronously triggers its phosphorescence due to the ordered molecular arrangement after molecular motion. The thermal effect is expected to be important for triggering efficient phosphorescence, and the photon input for providing a precise and non-invasive stimulus. Such sequential photo-thermo-phosphorescence conversion is anticipated to unlock a new stimulus-responsive phosphorescence material without chemicals invasion.
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