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Graphene Quantum Dot-MnO2 Nanosheet-Based Optical Sensing Platform: a Sensitive Fluorescence “Turn Off-On” Nanosensor for Glutathione Detection and Intracellular Imaging Xu Yan, Yang Song, Chengzhou Zhu, Junhua Song, Dan Du, Xingguang Su, and Yuehe Lin ACS Appl. Mater. Interfaces, Just Accepted Manuscript • DOI: 10.1021/acsami.6b05465 • Publication Date (Web): 05 Aug 2016 Downloaded from http://pubs.acs.org on August 5, 2016

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ACS Applied Materials & Interfaces

Graphene Quantum Dot-MnO2 Nanosheet-Based Optical Sensing Platform: a Sensitive Fluorescence “Turn

Off-On”

Nanosensor

for

Glutathione

Detection and Intracellular Imaging Xu Yan

, Yang Song2,‡, Chengzhou Zhu2, Junhua Song2, Dan Du*,1,3, Xingguang

1,2,‡

Su*,2 and Yuehe Lin*,1

1

School of Mechanical and Materials Engineering, Washington State University, Pullman, Washington 99164, United States

2

Department of Analytical Chemistry, College of Chemistry, Jilin University, Changchun, 130012, China

3

Key Laboratory of Pesticide and Chemical Biology of the Ministry of Education, P. R. China and College of Chemistry, Central China Normal University, Wuhan, 430079, P. R. China

ACS Paragon Plus Environment


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ABSTRACT Glutathione (GSH) monitoring has attracted extensive attention because it serves a vital role in human pathologies. Herein, a convenient fluorescence “turn off-on” nanosensor based on graphene quantum dots (GQDs)-manganese dioxide (MnO2) nanosheet has been designed for selective detection of GSH in living cells. The fluorescence intensity of GQDs can be quenched by MnO2 nanosheets via a fluorescence resonance energy transfer. However, GSH can reduce MnO2 nanosheets to Mn2+ cations and release GQDs, causing sufficient recovery of fluorescent signal. The MnO2 nanosheets serve as both fluorescence nanoquencher and GSH recognizer in the sensing platform. The sensing platform displayed a sensitive response to GSH in the range of 0.5-10 µmol L-1, with a detection limit of 150 nM. Furthermore, the chemical response of the GQDs-MnO2 nanoprobe exhibits high selectivity toward GSH over other electrolytes and biomolecules. Most importantly, the promising platform was successfully applied in monitoring the intracellular GSH in living cells, indicating its great potential to be used in disease diagnosis. Meanwhile, this GQDs-MnO2 platform is also generalizable and can be easily expanded to the detection and imaging of other reactive species in living cells. KEYWORDS: graphene quantum dots, manganese dioxide nanosheets, glutathione, fluorescence resonance energy transfer, intracellular imaging


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INTRODUCTION Glutathione (GSH), one of important non-protein thiol in eukaryotic cells, serves various indispensable functions including maintaining redox homeostasis status and defending against free radicals.1,2 Furthermore, it associates with the modulation of cell proliferation because it can protect the cells from oxidative stress and trap free radicals away from damaging DNA and RNA.3,4 More importantly, due to its significant effect on antioxidant aspect, GSH plays a vital role in the pathophysiology of a certain number of diseases, such as cancer, Alzheimer’s disease,5 human immunodeficiency virus (HIV),6 and diabetes.7 Therefore, sensitive intercellular GSH detection has attracted significant attention over past decades. Recently, several techniques have been proposed for detection of GSH such as high-performance liquid chromatography,8 electrochemistry,7 colorimetric assay,9 photoelectrochemistry,10 surface-enhanced raman scattering,11 and enzyme linked immunosorbent assay.12 Most of these techniques suffer from time-consuming, labor-intensive, sophisticated instrumentation, or high-cost biological reagents, which limits their practical applications.13 Considering the urgent metabolic functions of GSH for cellular homeostasis, establishing novel sensors is still desirable for sensitive detection of intercellular GSH level. Currently, tremendous studies have been made to develop fluorescent sensors for detection of various analytes14 and intracellular imaging.15 Fluorescent graphene quantum dots (GQDs), an exciting zero-dimensional nanomaterial with many attractive advantages including extraordinary optical, electronic, and biochemical properties,

such

as

size-dependent

tunable

photoluminescence,


excellent


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photoluminescence stability, low toxicity, and high biocompatibility.16-19 As a green substitute for the toxic transition metal quantum dots, GQDs have more promising applications in a broad range of areas such as cellular imaging,20 sensing17,21-25 and drug delivery.26 These attractive superiorities of GQDs make them as ideal chem/bio-sensors and bioimaging platforms. The combination of fluorescent probes and other functional nanomaterials in one micro matrix have been widely exploited in the construction of ideal sensors. The quenching effect induced by nanomaterials, such as graphene oxide,27,28 molybdenum sulfide,29 tungsten sulfide, gold nanoparticles30 and silver nanoparticles,31 has already been employed to develop fluorescence sensing platforms for sensitive detection of various analytes.32-34 Nonetheless, a critical limitation of developing these sensing platforms is the few reagents for switching the fluorescence of probes. Manganese dioxide (MnO2) nanosheets, with high specific surface area and superior light absorption capability, are promising fluorescence quenchers and have attracted significant attention in developing “turn off-on” fluorescence sensing platforms.35,36 Moreover, MnO2 nanosheets can be reduced to Mn2+, which may be friendly to the environment and human health.37,38 For example, Liu’s group first synthesized the MnO2 nanosheets conjugated with nanoparticles, yielding a promising fluorescence quencher for up-converted luminescence.39 Zhang et al. demonstrated a simple methods for the synthesis of MnO2 modified C3N4 nanosheets sandwich nanocomposites for GSH detection.40 However, relatively poor stability of MnO2-probe nanocomposites has limited these nanoprobes for GSH detection in vitro,


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due to the low solubility of bulk MnO2. Traditionally, MnO2 nanosheets synthesized through the top-down approach usually suffer the issues of high-cost, tedious, multistep processes for exfoliation and an additional separation step to obtain uniform nanosheets. Only one bottom-up strategy has been reported to prepare MnO2 nanosheets using oxidant as exfoliating agent.41 Thus, a practical challenge still remains to achieve convenient synthesis of uniform MnO2 nanosheets. Inspired by preceding work, we designed a sensitive and convenient optical sensing platform using ultrathin MnO2 nanosheets and GQDs as the energy donor-acceptor pairs (Scheme 1). Herein, we successfully synthesized GQDs by one-step hydrothermal treatment of graphene oxide (GO) in the ammine solution. In addition, the uniform MnO2 nanosheets were synthesized via a template-free, one step redox reaction between KMnO4 and ethanol. In the proposed sensing system, the ultrathin MnO2 nanosheets can quench the fluorescence (FL) intensity of GQDs through fluorescence resonance energy transfer (FRET). In addition to being efficient nanoquenchers for FL probes, MnO2 nanosheets can also serve as recognition units for GSH. GSH with reducing ability can trigger the decomposition of the MnO2 nanosheets which are selectively reduced into Mn2+, accompanying the subsequent recovery of FL intensity of GQDs. The introduction of MnO2 nanosheets not only improves the sensitivity but also specificity of response to GSH concentration, making it feasible for measuring the intracellular GSH concentration. The as-prepared nanosensor exhibits great potential in biological applications for GSH-associated disease monitoring and clinical diagnostics.



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EXPERIMENTAL SECTION Reagents and instruments. KMnO4, L-glutathione, and 4-morpholineethane sulfonic acid of analytical grade were purchased from Sigma-Aldrich Corporation. All reagents were used as received without further purification. The water used in this study had a good resistivity (> 18 MΩ cm-1). UV–vis spectra were recorded by a UV-2450 spectrometer. The FL spectra were measured by a Cary Eclipse fluorimeter. Cellular imaging was performed with a confocal laser fluorescence microscope (Leica SP8 Point Scanning Confocal). Transmission electron microscopy (TEM) images were carried out with a Philips CM200UT transmission electron microscope. Preparation of GQDs. GQDs were obtained by hydrothermal treatment of GO in ammonia solution. Typically, 30 mL GO dispersed water solution (1 mg mL-1) was mixed with 0.3 mL ammonia solution (28 wt %). Then, the mixed solution was transferred to Teflon lined autoclave and heated at 180 °C for 12 h. After cooling to room temperature, the black precipitated reduced graphene oxide (RGO) was removed. The light yellow supernatant was dialyzed for two days to remove excess ammonia. Finally, the nitrogen doped GQDs (N-GQDs) were obtained. Preparation of MnO2 Nanosheets. Ethanol (99%, 3.2 mL), sodium dodecyl benzene sulfonate (SDBS) (0.5 M, 1.6 mL), and H2SO4 (0.1 M, 0.16 mL) were mixed into 24 mL distilled water. The solution was heated to 90°C. KMnO4 solution (0.05 M, 0.32 mL) was slowly dropped to the mixed solution and keeped strring for 30 min to achieve red brown solution. The MnO2 nanosheets were purified rinsing


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with deionized water and ethanol three times. Then purified MnO2 was dispersed in DI water to further use. Detection Procedure. For the detection of GSH, different concentrations of GSH were mixed with 37.5 µg mL-1 MnO2 nanosheets and 200 µL purified GQDs solution in a series of 1.5 mL calibrated test tubes. Then, the mixtures were diluted to 1.5 mL with pH 6.5 PBS buffer (10 mmol L-1). The FL spectra were obtained after shaking for 6 min at room temperature. The in vitro imaging. Nanoprobe was dispersed into DMEM as stocking solution, then filtered and sterilized stocking solutions containing nanoprobe (0.5 µg µL-1) were added to the 6-well plate containing MCF-7 cultured by culture medium (2 mL). After incubation of cells with one or two hours in an incubator at 37˚C with CO2 (5%), the culture medium of cells was washed out by 1 × PBS for cell imaging. The bright and fluorescent fields of as-treated MCF-7 were observed with the 40× objective of confocal laser fluorescence microscopy.

RESULTS AND DISCUSSION Characterization of the GQDs and MnO2 nanosheets. To construct the sensing platform, GQDs were first prepared through the hydrothermal treatment method.42 The UV-vis absorption, FL excitation and emission spectroscopy were studied to investigate the optical properties of GQDs (Figure 1A). The GQDs show a weak shoulder peak around 330 nm, which may be originated from n-π* transitions of C=O band. The fluorescence excitation and emission spectra of GQDs exhibit obvious



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excitation peaks around 330 nm and 420 nm, respectively. The inset in Figure 1A shows the photographs of the GQDs under daylight and 365 nm UV irradiation. Figure 1B shows the transmission electron microscopy (TEM) image of GQDs. The size distribution of GQDs was reasonably uniform and the spherical particle size was approximately 8 nm. These results further demonstrated that GQDs with good water dispersibility were successfully prepared.42 Ultrathin MnO2 nanosheets, which were synthesized by KMnO4 reduction, were also characterized by UV-vis spectroscopy. As displayed in Figure 1C, the UV-vis absorption spectrum of MnO2 nanosheets expressed a broad absorption spectrum from 300 to 600 nm, which exhibited a characteristic absorption centered at 380 nm. Furthermore, ultrathin lamellar MnO2 nanosheets with occasional folds and wrinkles have been observed with TEM (Figure 1D), exhibiting a typical two dimensional morphology, and the average lateral dimension is estimated to be ～200 nm (Figure S1A). As shown in Figure S1B, X-ray photoelectron spectroscopy (XPS) of MnO2 nanosheets showed two peaks located at 642.0 and 653.9 eV, corresponding to Mn2p3/2 and Mn2p1/2 of MnO2, respectively. The energy separation of 11.9 eV is also consistent with previous reports.33 The Raman spectrum of the MnO2 nanosheets exhibited a peak at 653 cm-1, which was attributed to the Mn-O vibration (Figure S1C). Detection strategy of GSH. Scheme 1 illustrates the strategy of GQDs-MnO2 sensing platform for GSH detection. The basic principle of the GQDs-based GSH sensor includes the fluorescent quenching of GQDs by the MnO2 nanosheets and the decomposition of the MnO2 to Mn2+ by GSH. Primarily, as shown in Figure 2A,


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GQDs solution exhibits a maximum fluorescent signal at 420 nm (black curve). Compared to other signal sources, GQDs have been broadly applied in sensing systems due to their good biocompatibility, resistance against photobleaching and excellent photostability. The FL intensity of GQDs can be effectively quenched by MnO2 nanosheets (red curve). After adding GSH, MnO2 nanosheets can be effectively reduced to Mn2+ because of the special reaction between MnO2 nanosheets and GSH. As shown in equation (1),39,40 GSH was oxidized to produce glutathione disulfide. 2GSH + MnO2 + 2H+ → GSSG + Mn2+ + 2H2O

(1)

Due to the deposition of MnO2 nanosheets, the FL intensity of GQDs was distinctly recovered (blue curve). To investigate the feasibility of this proposed sensing system for GSH, the GQDs were firstly mixed with GSH. As shown in Figure 2B, different concentrations of GSH (from 0 to 200 µmol L-1) cannot induce obvious FL intensity changes to the GQDs system. Thus, the designed nanosensor can be used for GSH detection based on the FL recovery of GQDs. The proposed sensing platform consists of GQDs and MnO2 nanosheets, where GQDs function as fluorometric reporter, and MnO2 nanosheets serve bi-functionally as fluorescence quencher and GSH recognizer. The FL of GQDs can be efficiently quenched by MnO2 nanosheets through FRET.43,44 To clearly understand the quenching mechanism caused by MnO2 nanosheets, corresponding experiments were performed to study each of these effects. According to previous research, the absorption spectrum of MnO2 nanosheets overlaps well with the relative FL emission spectrum of GQDs (Figure S2), which is the indispensable factor for the generation



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of FRET.13,43 The TEM images of the MnO2-GQDs mixture is observed, which clearly display that the GQDs were absorbed on the surface of MnO2 nanosheets (Figure S3A). Furthermore, the FRET between MnO2 nanosheets and GQDs was further performed by FL lifetime measurements (Figure S3B). The lifetime of GQDs were obviously changed after adding MnO2 nanosheets. With the increasing of MnO2 nanosheets from 12.5 to 25.0 µg mL-1, the lifetime of GQDs became shorter, indicating that the dynamic quenching by FRET. To investigate the FRET between GQDs and MnO2, the quenching efficiency of the nanosensor was investigated by employing different concentrations of MnO2 nanosheets. As displayed in Figure 2C, a gradual decrease of the FL intensity of GQDs was observed with increasing concentration of MnO2 nanosheets in the range from 0 to 50 µg mL-1. Then, the FL remained stable with further increasing concentration of MnO2 up to 87.5 µg mL-1. An excellent linear relationship between quenching efficiency (FQ0-FQ)/FQ0 and MnO2 concentration was obtained with a regression coefficient of 0.990 (the concentration of MnO2 ranged from 0 to 37.5 µg mL-1) (Figure 2D). FQ and FQ0 are FL intensities in the presence and absence of MnO2 nanosheets, respectively. Therefore, the FL intensity of the GQDs could be modulated by the absorbance of MnO2 nanosheets via FRET. As a result, 37.5 µg mL-1 MnO2 nanosheets were selected for the subsequent experiments. Optimization of the sensing procedure. The sensitivity of the proposed sensor can be influenced by many factors, such as the pH and the reaction time, so the experimental conditions are optimized for determination of GSH. The effect of pH on


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the FL intensity ratio FR/FR0 of the GQDs-MnO2 probe in the presence of 10 µmol L-1 GSH was studied in Figure S4A. FR0 and FR were the FL intensity of the GQDs-MnO2 system in the absence and presence of GSH, respectively. The FL intensity ratio of the GQDs solution obviously enhanced with the change of pH value (5.5-6.5). Then gradual decreasing of intensity ratio was observed in the pH range from 6.5 to 8.5. At pH 6.5, the FL intensity ratio (FR/FR0) reached a maximum. Considering the above results, pH 6.5 PBS buffer (10 mmol L-1) was selected as the working pH for GSH detection. Incubation time was also a critical factor that could obvious influence the FL intensity of system, so time was studied in PBS buffer (pH 6.5) at room temperature (Figure S4B). When GSH was added into the GQDs-MnO2 system, the FL intensity enhanced immediately with the increasing reaction time and reached a balance in only 6 min. The working time was also verified by UV-vis absorption of MnO2 nanosheets in the presence of GSH (Figure S5). Thus, 6 min was chosen for sensitive detection of GSH in further experiments. Determination of GSH. To demonstrate the applicability of this developed“turn off-on” sensing platform for GSH detection, we investigated the response of approach to GSH. Under optimal conditions, the GQDs were mixed with 37.5 µg mL-1 of the MnO2 and different concentrations of GSH (0, 0.1, 0.5, 1.0, 2.0, 5.0, 10, 20, 50 and 00 µmol L-1) for 6 min at room temperature. The FL intensity of system was gradually restored with increasing concentration of GSH (Figure 3A, inset was the change trend of FL intensity ratio with different concentrations of GSH). Meanwhile, a good linear relationship over the range from 0.5 to 100 µmol L-1 with a correlation coefficient of



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0.992 was obtained (Figure 3B). The regression equation is: FR/FR0 =1.2745 + 0.3519 Log [GSH], µmol L-1. FR and FR0 are FL intensity in the presence and absence of GSH, respectively. The limit of detection (LOD) can reach 0.15 µmol L-1, which is calculated by the equation LOD = (3σ/s).45 In addition, the FL response of sensing probes with the use of 50 µg mL-1 MnO2 nanosheets was also studied. As shown in Figure S6, higher sensitivity was achieved when 37.5 µg mL-1 nanosheets was used. Moreover, our proposed method was compared with previous strategies for GSH detection in linear range and LOD (Table S1).6,9,39,40 This established GQDs-MnO2 system was comparable to or even better than most of the reported methods, indicating that the sensing platform has a good performance. Meanwhile, the GQDs-MnO2 system also showed much shorter analysis time than many previous sensors. Selectivity

of

GQDs-MnO2-based

fluorescence

toward

GSH.

Selective

recognition capability is a very significant character to investigate the performance of the FL sensing platform, especially for platforms with potential applications in intracellular detection and imaging. Therefore, to assess the specificity of the GQDs-MnO2 sensor for the detection of GSH, the influence of some common substance including inorganic salts (Na+, K+, Ca2+), metal ions (Mg2+ and Mn2+), amino acids (aspartic acid, tyrosine and glycine), glucose, fructose and proteins (bovine serum albumin (BSA), tyrosinase (TYR), acetyl cholinesterase (AChE), glucose oxidase (GOx), and trypsine (TRY)) was studied in aqueous solutions. As shown in Figure 4, after the addition of the above substances (100 µg mL-1 for protein


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and 500 µmol L-1 for others), the FL intensity of GQDs-MnO2 system remained nearly constant (FL intensity change less than 5%), and only after the addition of GSH, the FL intensity showed obvious recovery (Blank Column, Figure 4). The results indicated that the sensing platform showed the high selectivity to GSH. We further investigated the signal response of GQDs-MnO2 system to 50 µmol L-1 GSH in the presence of foreign substances. It could be seen from Figure 4 (Gray Column) that even in the presence of interference, the GQDs-MnO2 system still work the same for GSH recognition, indicating that the proposed GSH sensing system could ably resist interference from common electrolytes and biological species. Thus, the established nanosensor was suitable for selective detection of GSH in intracellular detection and imaging. In vitro GSH sensing. Cytotoxicity is an important issue needed to take into account for developing intracellular nanoprobe. The intrinsic cytotoxicity of MnO2 nanosheets, GQDs, and GQDs-MnO2 nanoprobes was evaluated by using MTT assay. MCF-7 cells were incubated with varieties of concentrations of MnO2 nanosheets, GQDs, and GQDs-MnO2 nanoprobes for 24 h. Figure S7 showed negligible loss of viability for MCF-7 cells after incubating with MnO2 nanosheets, GQDs and GQDs-MnO2 nanoprobe at concentrations below 40 µg mL-1, 60 µg mL-1 and 40 µg mL-1, respectively, which demonstrates that the GQDs and the MnO2 nanosheets have low cytotoxicity and favorable biocompatibility. These results suggested that the GQDs-MnO2 nanoprobe is an excellent candidate to be applied in intracellular detection.



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GSH serves various indispensable functions in biological systems. Moreover, the intracellular GSH concentration in cancer cells is up to millimolar range.46 As described above, we have demonstrated the effectiveness of GQDs-MnO2 nanoprobe system in sensing GSH based on “turn off-on” FL signal. Here we further investigate the capabilities of the as-prepared nanosensors for intracellular GSH monitoring in vitro. MCF-7 cells were incubated with the nanoprobe (40 µg mL-1) for various periods of time, whereas there was almost no FL signal for control MCF-7 cells only incubated with MnO2 (Figure 5). In addition, the FL intensity of MCF-7 cells was obviously enhanced with increasing the incubation time of nanoprobe because time-dependent delivered behavior into the cytoplasm and high level of GSH expression in MCF-7 cells. To evaluate the specific response of the nanoprobe to the changes of intracellular GSH concentration in vitro, the MCF-7 cells were pretreated with various thiol-reactive synthesis inhibitors, scavenger and enhancer, respectively. First, a series of control experiments were carried out that the cancer cells were incubated with GQDs only, GQDs with NMM and BSO for additional 2 and 4 h. Here, a continuously increased blue FL signal was achieved in cancer cells treated with GQDs only, indicating the successful transfer GQDs to cancer cells (Figure S8). As shown in Figure S8, the FL intensity of NMM or BSO-treated cells was similar to that of control, indicating that the FL intensity of GQDs was not mediated by GSH scavenger or inhibitors. Then, we evaluate the specific response of the nanoprobe to the changes of intracellular GSH concentration. It could be clearly shown from Figure S9 that obvious decrease of FL signals was observed in cancer cells with


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incubated with N-methylmaleimide (NMM, GSH scavenger: 500 µmol L-1) and buthionine sulfoximine (BSO, a GSH synthesis inhibitor: 200 µmol L-1) compared with the untreated one, due to decrease the intracellular GSH level. However, an FL enhancement for MCF-7 cells treated with α-lipolic acid (LPA, a GSH enhancer: 200 µmol L-1) was subsequently observed. These results implied that the GQDs-MnO2 nanoprobe is membrane-permeable and feasible for intracellular GSH monitoring in live cells.

CONCLUSIONS In conclusion, a convenient and sensitive FL nanosensor has been successfully developed through sequential assembly of GQDs-MnO2 nanoprobe for detecting GSH in living cell, which could be utilized to evaluate intracellular redox state. The GQDs served as the FL units and the MnO2 nanosheets were used as dual functions of nanoquenchers and recognizers in this sensing platform. The MnO2 nanosheets that quenched the FL intensity of GQDs through FRET can be selectively reduced to Mn2+ cations by GSH, along with the subsequent FL recovery of GQDs. Meanwhile, the nanosensors have a broad linear response to various concentration of GSH, making the nanosensor available to detect intracellular GSH level in cancer cells in vitro. As no further functionalization of MnO2 and GQDs are needed, the proposed platform shows excellent merits including cost-effectiveness, environmental-friendliness, and extremely fast and simple detection, suggesting this convenient method could work as an efficient sensing platform for GSH detection in biomedical applications.



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ASSOCIATED CONTENT Supporting Information TEM, XPR, Raman spectrum and UV-vis of the MnO2 nanosheets. Lifetime of GQDs with MnO2 nanosheets. Optimization conditions (pH and temperature). MTT assay of MnO2 nanosheets and GQDs. Confocal fluorescence microscope images of probe with NMM, BSO and LPA. Comparison between methods.

AUTHOR INFORMATION Corresponding Author *Email: [email protected] (D Du) *Email: [email protected] (YH Lin) *Email: [email protected] (XG Su) Notes The authors declare no competing financial interest.

Author Contributions ‡

X. Yan and Y. Song contributed equally to this work.

ACKNOWLEDGMENT This work was supported by WSU start up funding. We would like to thank the National Natural Science Foundation of China for financial support (no. 21575047) and China Scholarship Council (CSC) for the financial support.


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Scheme 1. Activation mechanism of the MnO2 nanosheet-GQDs nanoprobe for GSH detection in vitro.


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Figure 1. (A) The UV-vis absorption, excitation and emission spectra of GQDs; (B) The TEM image of GQDs; (C) UV−vis absorption of MnO2 nanosheets. The inset of A is photographs of the GQDs solution taken under visible light (left) and 365 nm UV light (right); (D) The TEM image of MnO2 nanosheets. The inset of C is a photograph of the MnO2 nanosheets solution taken under visible light.



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Figure 2. (A) The FL emission spectra of GQDs, GQDs/MnO2 and GQDs/MnO2/GSH. The inset of A is photographs of the corresponding color under 365nm UV lamp; (B) The influence of GSH concentration on the FL intensity of GQDs probe; (C) FL spectra of GQDs nanoprobe with different concentrations of MnO2 nanosheets. The concentrations of MnO2 nanosheets were 0, 2.5, 12.5, 18.75, 25, 37.5, 50.0, 62.5, 75.0 and 87.5 µg mL-1, respectively. The inset was the change trend of FL intensity ratio with different MnO2 nanosheets concentrations. (D) Relationship between quenching efficiency and the concentration of MnO2. FQ and FQ0 are FL intensities of GQDs in the presence and absence of MnO2 nanosheets, respectively.


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Figure 3. (A) FL spectra of GQDs-MnO2 sensing probe with different concentrations of GSH. The concentrations of GSH were 0, 0.1, 0.5, 1.0, 2.0, 5.0, 10.0, 20.0, 50.0 and 100 µmol L-1. The inset was the change trend of FL intensity ratio with different GSH concentrations. (B) Relationship between FL intensity ratio and the logarithm of GSH concentration. FR and FR0 are FL intensities of GQDs-MnO2 sensing probe in the presence and absence of GSH, respectively.



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Figure 4. The FL intensities of the GQDs-MnO2 system and GQDs-MnO2-GSH system in the presence of the interfering substances (100 µg mL-1 for protein and 500 µmol L-1 for others).


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Figure 5. Confocal FL microscopy images of MCF-7 cells incubated with MnO2 nanosheets and GQDs-MnO2 (40 µg mL-1): from top to bottom images represent MnO2 nanosheets and GQDs-MnO2 incubated for 1h and 2h.



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Table of Contents Graphic 254x190mm (96 x 96 DPI)


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Nanosheet Based Optical Sensing Platform: A ... - ACS Publications

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