Applying Carbon Dots-Metal Ions Ensembles as a Multichannel

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Applying Carbon Dots-Metal Ions Ensembles as a Multichannel Fluorescent Sensor Array: Detection and Discrimination of Phosphate Anions Shan Sun, Kai Jiang, Sihua Qian, Yuhui Wang, and Hengwei Lin Anal. Chem., Just Accepted Manuscript • Publication Date (Web): 28 Apr 2017 Downloaded from http://pubs.acs.org on April 29, 2017

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Analytical Chemistry

Applying Carbon Dots-Metal Ions Ensembles as a Multichannel Fluorescent Sensor Array: Detection and Discrimination of Phosphate Anions

Shan Sun,†,‡ Kai Jiang,† Sihua Qian,† Yuhui Wang,† and Hengwei Lin*,†

†

Key Laboratory of Graphene Technologies and Applications of Zhejiang Province, Ningbo

Institute of Materials Technology & Engineering (NIMTE), Chinese Academy of Sciences (CAS), Ningbo 315201, P. R. China ‡

University of Chinese Academy of Sciences, Beijing, 100049, P. R. China

*Email: [email protected]; Tel: +86-574-86685130; Fax: +86-574-86685163

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Analytical Chemistry 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

ABSTRACT Fluorescent carbon dots (CDs) are attracting much attention in sensing recently thanks to their superior optical properties and abundant surface functional groups. To take further advantages of these unique features, CDs are considered to be possible for facilely fabricating multichannel sensor arrays. As a proof-of-concept research, CDs-metal ions ensembles are screened and designed as a triple-channel fluorescent sensor array in this study for the identification of various phosphate anions (e.g. ATP, ADP, AMP, PPi, and Pi) for the first time. Further studies reveal that the selected three metal ions (i.e. Ce3+, Fe3+ and Cu2+) could induce aggregation of the CDs, resulting in quenching of their fluorescence. However, disaggregation or further aggregation of the CDs-metal ions ensembles occurs with the addition of phosphate anions. Consequently, fluorescence of the CDs is recovering or further quenching. On account of various numbers of phosphate group and steric hindrance effects of phosphate anions, their affinities to the sensor array can be distinguished through fluorescence changes of the CDs-metal ions ensembles. By means of statistical analysis methods, the as-developed array is shown excellent capabilities in the detection and discrimination of phosphate anions. Furthermore, practicability of the sensor array is validated by the successful identification of phosphates in serum and blind samples. Compared to previous reports, the as-developed multichannel sensor array manifests numerous advantages, such as simple fabrication process, flexible adjusting detection ranges, and possible extension to other analytes having similar chemical structures or properties.


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INTRODUCTION As a newly emerged type of fluorescence (FL) nanomaterials, carbon dots (CDs) have inspired intense research efforts in recent years.1-4 Comparing with traditional organic FL dyes and semiconductor quantum dots, CDs hold many merits including facile preparation, superior optical properties, easy surface functionalization, chemical inertness, low toxicity and good biocompatibility. Hence, CDs are demonstrated many potential applications, such as photocatalysis,5,6 optoelectronics,7-9 sensing,10 bioimaging,11-13 and theranostics.14,15 Among these fascinating applications, the use of CDs in sensing is one of the most promising fields thanks to their superior optical properties and possessing abundant surface functional groups. To take further advantages of these unique features, CDs are considered to be possible in facilely fabricating multichannel sensor arrays, but which had only been very little exploited.16 The array sensing technique has shown numerous advantages, such as accuracy, diversity and capacities in the simultaneous detection and discrimination of chemically structures or properties similar analytes, thus receiving much attention and broad applications in recent years.17-25 Based on cross-responsive sensing elements instead of specific receptors, sensor arrays could produce composite responses unique to one analyte, which is similar to the mammalian gustatory or olfactory systems.26-29 It’s well-known that phosphate anions play crucial roles in physiological processes concerned with phosphorylation, maintenance of phosphate homeostasis, nutrition balance, and even some severe diseases.30,31 Besides, the industrial and environmental effects of phosphate anions are also enormous because of their broad usage as fire retardants, waste water purifying agents, and fertilizers.30 Therefore, great efforts have been devoted to developing sensitive and selective methods for the detection of phosphate anions in the past decades, such as fluorometry,32,33

spectrophotometry,34

chromatography,35

enzymatic

biosensors,36,37

electrochemical analysis,38 and so on.39 All these methods, however, require sophisticated instrumentation and/or time-consuming testing process.40 Moreover, selectivity of the reported methods is usually problematic with the presence of other competing phosphates, thus seriously limiting their applicability in real conditions. To overcome these drawbacks, array sensing technique might be one of the choices. But, to the best of our knowledge, only one sensor array-based method had been reported for the detection and discrimination of various phosphate anions till today.41 Nevertheless, this reported array is difficulty in fabrication due to requiring ACS Paragon Plus Environment


complicated synthesis of sensing units. Therefore, it is of great importance and interests to develop new sensor systems that can identify and differentiate phosphate anions simultaneously. Given the significance and difficulties in the recognition of phosphate anions and verify feasibility of the above proposed CDs-based multichannel sensor array strategy, herein, CDs-metal ions ensembles are designed as a triple-channel FL sensor array for the detection and discrimination of various phosphate anions (e.g. ATP, ADP, AMP, PPi, and Pi). Owning to covering abundant functional groups on the surface of CDs and which making them possible to interact with metal ions, Ce3+, Fe3+ and Cu2+ are screened and selected to coordinate with the CDs, resulting in quenching their FL due to aggregation. With the addition of phosphate anions, disaggregation or further aggregation of CDs-metal ions ensembles occur depending on the used metal ions (i.e. disaggregation for the Fe3+/Cu2+-CDs ensembles and further aggregation for the Ce3+-CDs ensembles, respectively), thus inducing FL recovering or further quenching. Based on the distinguished FL changes of the CDs, these phosphate anions can be well identified and differentiated. Moreover, quantitative detection of ATP using this array and its potential applications for the identification of phosphate anions in complex media (e.g. blood serum) and blind samples were also demonstrated. As far as we know, this is the first example to fabricate CDs-based sensor array and achieve simultaneous detection of a variety of phosphate anions. Importantly, such a design strategy is believed to be able to extend to other analytes having similar chemical structures or properties.

EXPERIMENTAL SECTION Reagents and materials All chemicals from commercial sources are of analytical grade. Citric acid, Ce(NO3)3·6H2O, N-(2-hydroxyethyl)piperazine-N’-(2-ethanesulfonic acid) (HEPES), sodium acetate (NaAc), adenosine 5’-triphosphate disodium salt (ATP), adenosine 5’-diphosphate sodium salt (ADP), adenosine 5’-monophosphate sodium salt (AMP), sodium tripolyphosphate (PPi), sodium phosphate tribasic dodecahydrate (Pi), sodium sulfate anhydrous, calcium nitrate tetrahydrate, L-cysteine (Cys), L-threonine (Thr) and L-tryptophan (Try) were obtained from Aladdin. Formamide, acetone, methanol, acetic acid (HAc), potassium chloride, and glycine (Gly) were purchased from Sinopharm. Cu(ClO4)2, Fe(ClO4)3 and tris(hydroxymethyl) amino methane (Tris) were from J&K Scientific Ltd. Fetal bovine serum (FBS) was purchased from BioInd. ACS Paragon Plus Environment

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Instrumentation MDS-6G microwave chemical reactor (SMART, Shanghai SINEO Microwave Chemistry Technology) was used to synthesize carbon dots. Characterizations of carbon dots were performed by the following instruments. UV-Vis absorption spectra were recorded on a PERSEE T10CS UV-Vis spectrophotometer. Fourier transform infrared (FT-IR) spectra were obtained on a Nicolet 6700 FT-IR spectrometer. Transmission electron microscopy (TEM) observations were performed on a Tecnai F20 microscope. Atomic force microscope (AFM) measurements were carried out with Veeco Dimension 3100V. X-ray photoelectron spectroscopy (XPS) spectra were performed on an AXIS UL TRA DLD spectrograph with Al/Kα as the source. For all sensing processes, the fluorescence signals were collected from Hitachi F-4600 spectrophotometer at ambient conditions. Preparation of the carbon dots The preparation of the carbon dots was referenced to our previous study.42 Briefly, 50 mL of 5% (weight) citric acid formamide solution was transferred into a Teflon-lined autoclave and experienced two heating steps in the microwave chemical reactor. Firstly, heated at 160 ºC (400 W) for one hour, and then keeping at 120 ºC for another hour. After cooling the autoclave to room temperature naturally, the viscous liquid was aged in 250 mL acetone overnight in a refrigerator (−20 °C). The precipitation was then washed with acetone and 10% methanol/acetone three times, respectively (50 mL solvent being used for each washing step). Subsequently, the residue was re-dispersed in methanol and filtered through 0.22 μm membrane to remove large particles. Finally, the purified CDs were obtained by evaporating off methanol and further dried in vacuum oven and stored as a deep red powder. Sensing procedure Working solutions for fabricating the triple-channel sensor array: 20 μM Ce3+ in Tris (10 mM, pH 7.0), 50 μM Fe3+ in HAc-NaAc (10 mM, pH 5.0), and 10 μM Cu2+ in HEPES (10 mM, pH 7.4) were incubated with the CDs (3 μg/mL) for 10 min before the detection process. FL intensity changes of the working solutions were recorded in the absence and presence of desired concentrations of phosphate anions (i.e. ATP, ADP, AMP, PPi, and Pi). The experiments were performed in quintuplicate to investigate the reproducibility and stability of this system. Then, the sensor array’s selectivity was investigated through adding diverse interfering ACS Paragon Plus Environment


substances and measured in triplicate. Incubation time effects were measured in each step to make sure the validity of the recorded data. In addition, F-F0 was applied to show the FL response of the array to analytes, where F and F0 represent the array’s normalized FL intensities in the presence and absence of phosphate anions, respectively. Finally, principle component analysis (PCA) and hierarchicalcluster analysis (HCA) were performed through Multi-Variate Statistical Package (MVSPv.3.1, Kovach Computing). Analysis of real sample Phosphates detection in the presence of fetal bovine serum (FBS) was taken as an example to test the practicality of the as-developed array for real sample. FBS was firstly incubated with the working solution for 10 min (containing 0.1% of FBS), followed by adding different phosphate anions. The analysis process was the same as the above described sensing procedure. Blind sample test A total of 20 samples including 15 simulant challenges and 5 blanks were analyzed in the blind trial. Samples contain one of the phosphates in deionized water with concentrations between 1 to 10 μM (above the limit of detection). Three replicate samples were prepared for each kind of phosphates. The results were analyzed using the triple-channel FL recognition patterns to identify the phosphates through comparing the results with the trained “fingerprint maps”.21,43-46 RESULTS AND DISCUSSION Preparation and Characterizations of the CDs The CDs used for construction of the triple-channel FL sensor array in this study was prepared according to our previous study.42 The morphology, composition, and photophysical properties characterizations of the CDs including TEM, AFM, UV–Vis absorption, FL emission and excitation, XPS, and FT-IR were thoroughly performed (Figure S1-S3 in Supporting Information (SI)). All these data are closely similar as that of our previous report, indicating successfully obtaining the CDs. Specifically, the CDs are identified covering abundant surface functional groups on their surface (e.g. −NH2, −OH, and −COOH) based on the XPS C1s and FT-IR spectra analysis (Figure S2 and S3 in SI). These functional groups make the as-prepared CDs possible to chelate with various metal ions.


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Design and fabrication of the sensor array It was reported that metal ions could trigger aggregation of CDs to form chemosensing ensembles. 47,48

Thus, based on the competing interaction of metal ions with CDs and analytes, a

multichannel sensor array could possibly be fabricated by employing a series of appropriate CDs-metal ions ensembles by monitoring the FL changes of the CDs. Given the significance and difficulties in the detection and discrimination of phosphate anions, and as well as their unique affinity to certain metal ions, a triple-channel sensor array was attempted to fabricate in this study. To fulfill such a design, FL responses of the as-prepared CDs to a variety of metal ions (i.e. Ce3+, Fe3+, Cr3+, Zn2+, Fe2+, Cu2+, Hg2+, Ag+, and Pb2+) under different conditions (i.e. different buffers and pH) were thoroughly assessed. The results indicated that FL of the CDs was quenched in different degrees with introduction most of these metal ions (Table S1 in SI ), probably due to aggregations of the CDs.49-51 However, considering the potential toxicity, FL responding extents, and possible coordination with phosphates, three metal ions under specific conditions [i.e. Ce3+ in Tris (10 mM, pH 7.0), Fe3+ in HAc-NaAc (10 mM, pH 5.0), and Cu2+ in HEPES (10 mM, pH 7.4)] were selected to fabricate the CDs-metal ions ensembles based triple-channel sensor array. On account of different phosphate group numbers and steric hindrance effect of the phosphates (i.e. ATP, ADP, AMP, PPi, and Pi), their affinities to metal ions can be distinguished from the competitions with surface functional groups of the CDs, resulting in discriminative FL signal changes of the array and thus realizing identification of various phosphates. Such a design and fabrication process for the identification of different phosphate anions can be simply illustrated in Scheme 1. It’s worthy to note that this sensor array is facile and cost-effective in fabrication due to no tedious synthesis of multiple probes being needed.

Scheme 1. Schematic illustration of the design and fabrication process of the triple-channel FL sensor array for the detection and discrimination of phosphate anions.



Optimization of the detection conditions In order to obtain the best sensing performance, detection conditions were further optimized. Since the basic principle for fabricating the array is depending on different affinities between phosphates and metal ions, the applied concentrations of metal ions for making CDs-metal ions ensembles are critical. Thus, titration experiments of Ce3+, Fe3+ and Cu2+ against FL responses of CDs in their corresponding buffer conditions were carried out. From the results shown in Figure S4 (SI), about 20 μM Ce3+, 50 μM Fe3+, and 10 μM Cu2+ were observed to obtain nearly the maximum FL responses. Moreover, the incubation times for different steps along the sensing process were also investigated. Figure S5A (SI) shows that only two minutes needed to reach stable FL quenching responses with the addition of the three metal ions. Figure S5B-5D (SI) exhibit that the FL intensities of CDs-metal ions ensembles level off to saturations after ten minutes of incubation with the addition of phosphate anions. According to these results, 20 μM Ce3+, 50 μM Fe3+, and 10 μM Cu2+ with 10 min incubation time were finally chosen as the optimal detection conditions. Note that FL intensity changes of the CDs with direct addition of phosphate anions were measured as controls, and nearly no responses were observed (Figure S6 in SI), indicating negligible effects of phosphates to FL of the CDs. Detection mechanism of the sensor array In order to determine the possible mechanisms of the sensor array, TEM tests were performed for the CDs-metal ions ensembles in the absence and presence of the phosphate anions. It is clearly shown that the CDs are well dispersed (Figure S1A in SI), but aggregated to diverse extents in the presence of the metal ions (Figure 1A, 1D, and 1G). Among them, Cu2+ exhibits the strongest quenching ability to the FL of CDs, followed by Fe3+ and Ce3+. These results are in good accordance with the aggregation extents presented in TEM images. Then, the morphology changes of CDs-metal ions ensembles after the addition of phosphates were examined. Interestingly, the TEM tests reveal that more intense aggregation occurring for the CDs-Ce3+ensembles with the addition of phosphates (Figure 1B and Figure S7B in SI as two typical examples), while different degrees of disaggregation for CDs-Fe3+ and CDs-Cu2+ ensembles (e.g. Figure 1E and 1H, and Figure S7D and S7F in SI). In fact, these aggregation or disaggregation changes can also be directly observed by the naked eye (Figure 1C, 1F, and 1I) if higher concentrations of metal ions and phosphates were added. The opposite phenomena of


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CDs-metal ions ensembles with addition of phosphates (i.e. disaggregation or more intense aggregation) could be explained as follows: i) Fe3+ and Cu2+ should have stronger coordination capability to phosphate anions than to the functional groups on CDs, and thus they are replaced from the CDs with the addition of phosphates, consequently inducing CDs’ disaggregation and FL recovering; and ii) the observed further aggregation and quenching of the CDs with the addition of phosphates to the CDs-Ce3+ ensemble might be ascribed to the higher coordination number of Ce3+ and its similar coordination capability with phosphates and functional groups on the CDs, and thus Ce3+ and phosphates could behave as bridges to connect more CDs together.

A

D

G

B

E

H

C

CDs + Ce3+

F

CDs + Fe3+

CDs + Cu2+

I

Figure 1. TEM images of the CDs with the addition of Ce3+ (A), Fe3+ (D), and Cu2+ (G), respectively; TEM images of the CDs-Ce3+ ensemble with the addition of Pi (B), CDs-Fe3+ ensemble with the addition of ATP (E), and CDs-Cu2+ ensemble with the addition of ATP (H); photographs of the CDs with the addition of Ce3+ and then add Pi (C), the CDs with the addition of Fe3+ and then add ATP (F), and the CDs with the addition of Cu2+ and then add ATP (I).

Responses of the array to phosphate anions and interfering substances To examine the identification capability of the as-fabricated array, its FL responses to a variety of phosphate ions (e.g. ATP, AMP, ADP, PPi, and Pi) were firstly recorded. As shown in Figure 2A,



the array exhibits unique triple-channel FL recognition patterns (or “fingerprint maps”) to each of these phosphates (all at 10 μM). Then, principal component analysis (PCA), a statistical procedure that uses an orthogonal transformation to convert a set of observations of possibly correlate variables into a set of values of linearly uncorrelated variables, was applied to these triple-channel FL responses. It can be seen from Figure 2B, all of these investigated phosphate anions are separated exactly from each other with no errors or misclassifications in quintuplicate. Meanwhile, the selectivity and anti-interference capability of the developed array were investigated as well. No obvious array responses were observed with the addition of Ac−, Cl−, SO42−, K+, Na+, and Ca2+ even at a relative high concentration (50 μM). In addition, amino acids, such as Gly, Try, Thr, and Cys (10 μM), were also introduced to the sensor platform as interfering substances. Although these amino acids could also induce array’s responses, they are clearly separated from the phosphates. To further evaluate the discriminative capability of this sensing array, another set of concentration of phosphates (all at 5 μM) were also examined. Again, each phosphate anion induces a specific array response, and is clustered into separated group by the PCA (Figure S8B in SI). Besides, hierarchical cluster analysis (HCA) was also performed to provide further evidence for the array’s recognition capability for phosphate anions. Similarly to the PCA, phosphates and other interfering substances are classified into different clustering groups, displaying clear discrimination of these phosphate anions and as well as from the interferants (Figure 2C and Figure S8C in SI). These results clearly demonstrate that the as-fabricated sensor array is of high specificity and differentiation capability for phosphate anions. A Normalized（F-F0）

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Amino acid Metal ions， anions and control

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Thr Try Cys Gly Control Na2SO4 KCl Ca2+ NaAc AMP Pi ADP PPi ATP

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Figure 2. (A) FL responses of the triple-channel sensor array to five phosphate anions (all at 10 μM), in which F and F0 represent the array’s FL intensities of the CDs-metal ions ensembles in the presence and absence of phosphate anions, respectively; (B) principal component analysis (PCA) plot for the discrimination of the phosphate anions and other interfering substances based on the FL signal changes of the sensor array; (C) hierarchical cluster analysis (HCA) plot for the discrimination of the phosphate anions and other interfering substances based on the FL signal changes of the sensor array.

Since the concentrations of metal ions not only affect the FL quenching efficiencies to the CDs, but also influence the amount of phosphate anions to coordinate with them, therefore, the detection ranges of the array for the phosphates could be adjusted by altering the concentration ratios in making the CDs-metal ions ensembles. To clarify this issue, the concentration of Ce3+ was increased from 20 to 50 μM to make the corresponding CDs-Ce3+ ensemble, and then the array was applied to higher concentration of phosphates (all at 50 μM) and corresponding FL responses were recorded (Figure S9A in SI). Similarly as above, phosphate anions, amino acids and other interfering substances are firstly classified principally into three main groups, and further the phosphates are separated from each other clearly (Figure S9B,C in SI). Note that all of ACS Paragon Plus Environment


the investigated amino acids can also be discriminated against one another apparently. These results demonstrate good expandability of the array in the ranges of application, such as detection concentrations of the phosphates and types of analytes. In addition, it’s necessary to point out that a contrary FL enhancement rather than quenching of CDs-Ce3+ ensemble was observed with the addition of ATP herein (Figure S9A in SI). This phenomenon could be explained as follows: when a low concentration of ATP was added, FL of the CDs-Ce3+ ensemble was further quenched due to formation of large aggregates; but a relatively high concentrations of ATP would induce disaggregation of the CDs-Ce3+ ensemble and release free CDs, resulting in the FL recovering (Figure S10 in SI). Quantitative detection of ATP To illustrate the quantitative detection application of the as-developed array, quantification of ATP is taken as an example due to its unique functions as the extracellular signaling agent in biological process and energy source for biological reaction.52 In order to ensure the presented system can be used for quantification, the FL responses (illustrated by the total Euclidean distances (EDs), i.e. square root of the sums of the square of the normalized (F-F0) values) induced by ATP at different concentrations were evaluated. As shown in Figure 3, an obvious increase of the EDs values was observed as the concentration of ATP increased from 0.5 to 10 μM. Moreover, the inset of Figure 3 clearly reveals a linear relationship between EDs values and the concentrations of ATP in the range from 0.5 to 6 μM. Based on the linear fitting parameters, a limit of detection was then calculated to be 0.11 μM using the well-known 3σ (signal-to-noise) criteria.53 0.4 0.3 0.2

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R2=0.9995

EDs

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Figure 3. The total Euclidean distances (EDs) of the array plotted versus different concentrations of ATP. Inset: the linear relationship in the concentration of ATP from 0.5 to 6 μM (error bars illustrate three parallel measurements). ACS Paragon Plus Environment

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Real sample assay Finally, the potential practicalities of the triple-channel FL sensor array for the identification of phosphate anions were evaluated in blood serum samples. Blood serum is known containing not only proteins and electrolytes, but also phosphates and various carboxylate ions,41 which is likely to cause a unique response pattern of this array. Similarly, the triple-channel PL responses of the array to serum and serum in the presence of phosphate anions (all at 10 μM) were measured and then analyzed using PCA and HCA. As expected, the serum itself created a unique array’s response, and the five phosphate anions distinguished from each other (Figure 4). Note that similar array responses were observed herein for serum itself and serum plus Pi (probably due to containing complicated components in serum such as proteins, electrolytes, and even phosphates), but discrimination of them is still no errors by HCA (Figure 4B). Moreover, even lower concentration of phosphates (5 μM) in serum can also be differentiated by this array (Figure S11 in SI), revealing potential applications of the as-developed FL sensor array for phosphate anions in real samples. a)

PC 2 (14.7%)

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Figure 4. Principal component analysis (PCA) (a), and hierarchical cluster analysis (HCA) (b) plots for the discrimination of five phosphate anions (all at 10 μM) in the presence of fetal bovine serum based on the FL signal changes of the as-developed triple-channel sensor array. ACS Paragon Plus Environment


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Blind laboratory trial To further demonstrate the practicalities of the as-developed triple-channel sensor array, blind laboratory tests were conducted. Of the 20 blind samples, 15 were used as challenges, which contain one of the five kinds of phosphate anions, and 5 were blanks. Their FL recognition patterns were obtained by the same approach (Figure S12 in SI). All of the positive samples (15 of 15) were correctly identified, yielding no false negative. All of the five blank samples were also unanimously recognized, yielding 0% false positives (Table S2 in SI). CONCLUSIONS In summary, in order to extend the application ranges of CDs in sensing field, a multichannel sensing strategy was proposed in this study for the first time. As a proof-of-concept research, a triple-channel FL sensor array based on the CDs-metal ions ensembles was designed and constructed. The as-fabricated array was demonstrated to be excellent not only in the detection and discrimination of five phosphate anions (e.g. ATP, ADP, AMP, PPi, and Pi), but also in quantitative detection. Further studies revealed that disaggregation or further aggregation of the CDs-metal ions ensembles occurred with the addition of phosphates depending on the nature of metal ions used for making the ensembles. Moreover, this array method as well show high sensitivity, good anti-interference and potentials in complex media like blood serum. The high recognition accuracy of this sensor array to blind samples also validated its practicalities. Note that this research is regarded as a preliminary exploration of fabricating CDs-based sensor array systems. The identification of other chemically/property similar analytes (e.g. amino acids, peptides, and proteins) is believed to be possible as well through adjusting the types of CDs-metal ions ensembles and the detection conditions. ASSOCIATED CONTENT Supporting Information The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.analchem.xxxxxxx. The TEM, AFM, XPS, FT-IR, and optical characterizations of the CDs, supplementary figures and tables (PDF). AUTHOR INFORMATION


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Corresponding Author [email protected] Notes The authors declare no competing financial interest. ACKNOWLEDGMENTS This work was financially supported by the Natural Science Foundation of China (21607160), the Zhejiang Provincial Natural Science Foundation (LY16B050005), and Ningbo Science and Technology Bureau (2014A610195, 2014B82010, 2015A610277 and 2016C50009). REFERENCES 1.

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