English

• 0.   Introduction

  Diabetes mellitus, a chronic disease, is caused by insufficient insulin secretion in the body or inefficient use of insulin by cells^[1-2]. This disease is characterized by high blood sugar, which can induce a variety of chronic complications, such as blindness, coronary heart disease, cerebrovascular disease, and diabetic nephropathy^[3-4]. Nowadays, there were approximately 463 million people suffer from diabetes in the world. Unfortunately, the number of diabetic patients has continued to increase for over 50 years according to the latest report from International Diabetes Federation^[5]. In the future, it is predicted that there will be 700 million diabetic patients worldwide by 2045, which will be seriously threatened human health^[6]. Given the definitive functional relationship between human health and glucose, the development of a glucose sensor with high sensitivity, wide linearity, low cost, and strong antiinterference would be of great significance for assisting the clinical assessment of diabetes.

  At present, there are various technologies that were used for glucose detection, such as colorimetry^[7], Raman spectroscopy^[8], fluorescence spectroscopy^[9], photoelectrochemical measurement^[10], gas chromatography-mass spectrometry^[11], and electrochemical method^[12]. Among these methods, electrochemical sensors have received widespread attention in the multiplex biological matrix because of their high discernibility, fast response, low detection limit, simplicity, as well as low cost^[13-14]. Generally, the most widely-used electrochemical glucose sensors are enzyme-based sensors^[15]. However, enzymatic electrochemical sensors have some poor stability, complicated immobilization procedures, critical operating situation, and high cost that limit their practical application in glucose detection^[16-17]. Because of the above-mentioned drawbacks, non-enzyme electrochemical glucose sensors have become a new research hotspot. For non-enzyme electrochemical glucose sensors, the key factor is electrode materials. From this perspective, the foremost thing is to find novel electroactivity materials, which could catalyze glucose oxidation with fast kinetics and lower overpotential.

  In this direction, various nanomaterials have been studied for fabricating non-enzyme glucose sensors, such as carbon nanomaterials^[18], metal-organic frameworks nanomaterials^[19], and quantum dots^[20], metal and metal oxide nanomaterials^[21]. Thereinto, Pt-based materials expressed prominent catalytic activity for the oxidation of glucose and were widely used to construct an electrochemical interface for non-enzyme-sensors^[22]. For instance, Guo et al. used ultrasonic electrodeposition to deposit Pt nanoparticles onto gold electrodes, which showed good catalytic performance in a neutral solution with a linear range of 1-16 mmol·L^-1 and a detection limit of 48 µmol·L^{-1 [23]}. However, the intermediate products produced in the process of glucose oxidation easily adhere to the Pt nanomaterials, which leads to the passivation or poisoning of the interface and reduces the electrocatalytic activity subsequently^[24]. To settle down this limitation, developing bimetallic nanomaterials is considered to be an effective way. Gold (Au) nanomaterial is another interesting and attractive material due to the merits of high surface reactivity, good solubility, and excellent bioactivity^[25]. Meanwhile, Au also has strong anti-interference, which is attributed to its full-filled 3D orbit. In previously published reports, bimetallic Pt/Au showed excellent electrocatalytic activity toward glucose and high tolerance to toxicity. However, the high price and large size of electrode substrate are inconvenient for the in-vivo detection and limited their application.

  Acupuncture is a kind of traditional Chinese medicine treatment technique, which can relieve pain and promote blood flow by piercing a needle into the human body^[26]. Acupuncture needles (ANs) are made of stainless steel, gold, silver, etc. Among these materials, the stainless steel AN is cheap and commonly used. Because ANs are made of metal with high conductivity, which provides their potential applications as electrochemical electrodes. Currently, relative reports have confirmed that ANs can be used as electrodes for the detection of nitrate^[27], lead^[28], rutin^[29], ascorbic acid^[30], dopamine^[31], hydrogen peroxide released from living cells^[32], etc. Compared with the common glassy carbon electrode, the AN electrode only needs to be cleaned rather than polished before electrochemical experiments. What′s more, it has been reported that the unique needle-shaped structure provides a larger surface area for the modifier and improves electrode performance^[33-34]. In a word, stainless steel AN has the advantages of low price, simple to operate, acicular structure, and high specific surface area. However, the detection of glucose based on ANs has not been reported so far, although it is very significate and strong practicable, which should be anastomosed with traditional medicine acupuncture.

  In this work, a bimetallic Pt/Au nanocatalyst was modified onto stainless steel AN tip by electrodeposition, and the interface possessed the intrinsic capability for electrochemical non-enzyme sensing glucose effectively. Electrodeposition is a feasible method to synthesize metal, in which the size and morphology of alloy can be adjusted by applied potential, deposition time, and concentration. The catalytic performance of the non-enzyme electrochemical sensor (Pt/Au/AN) for glucose oxidation was systematically studied. Compared with Pt/AN and Au/AN electrodes, Pt/Au/AN electrodes significantly improved the electrocatalytic performance of glucose oxidation. Meanwhile, Pt/Au/AN electrode possesses not only simple preparation and low price, but also high selectivity, sensitivity, and a wide linear range for detecting glucose.

  1.   Experimental

  1.1   Reagents

  Gold chloride trihydrate (HAuCl₄·3H₂O), chloroplatinic acid hexahydrate (H₂PtCl₆·6H₂O), glucose, sodium hydroxide (NaOH), uric acid (UA, >98%), ascorbic acid (AA, ≥99%) were purchased from Aladdin Reagent Co., Ltd. Sulfuric acid (H₂SO₄) and absolute ethanol were obtained from Sinopharm Chemical Reagent Co., Ltd. Stainless steel ANs (size: 0.25 mm× 40 mm) were purchased from Suzhou Medical Appliance Factory Co., Ltd. All reagents were analytical grade and the aqueous solution was prepared using 18.25 MΩ·cm^-1 ultrapure water throughout the experiment. Human serum samples were taken from Hangzhou Cancer Hospital, affiliated with Zhejiang University, China.

  1.2   Apparatus

  The morphology of the prepared electrode was characterized by Gemini scanning electron microscope 300 (SEM, ZEISS, Germany) operating at 1, 2, 3, or 5 kV. On the CHI660 electrochemical workstation (Chenhua, Shanghai, China), the conventional three-electrode mode was used for electrochemical measurement. The modified electrode, saturated calomel electrode (SCE), and Pt wire electrode were used as the working electrode, the reference electrode, and the counter electrode, respectively.

  1.3   Preparation of the modified electrode

  Scheme 1 illustrates the preparation procedure of Pt/Au/AN electrode for the determination of glucose. As shown in Scheme 1A, Pt/Au/AN electrode was fabricated by a two-step sequential electrodeposition strategy. Firstly, the AN was ultrasonically cleaned in absolute ethanol and ultrapure water for 5 min and then dried with nitrogen. Subsequently, Au/AN electrode was achieved by depositing gold nanomaterials onto AN in the range of 1.5 mm from the tip via applying an accumulation potential of -8.0 V for 180 s in 0.75 mmol·L^-1 HAuCl₄ solutions (HAuCl₄·3H₂O was dissolved in 0.2 mol·L^-1 H₂SO₄). Finally, Pt/Au/AN electrode was obtained by modifying Pt nanomaterials onto Au/AN surface via performing an accumulation potential of -0.8 V for 300 s in 1.25 mmol·L^-1 H₂PtCl₆ solutions (H₂PtCl₆·6H₂O was dissolved in 0.4 mol·L^-1 H₂SO₄). Pt/AN and Au/AN electrodes were used as the control, which were prepared by a one-step electrodeposition procedure via immersing the bare AN electrode into 1.25 mmol·L^-1 H₂PtCl₆ solutions or 0.75 mmol·L^-1 HAuCl₄ solutions, and then performing an electrodeposition procedure, respectively.

  Scheme 1

  

  Scheme 1.  (A) Fabrication procedure of Pt/Au/AN electrode; (B, C) Mechanism of glucose oxidation on Pt/Au/AN electrode

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  1.4   Electrochemical measurements

  Firstly, cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were used to characterize the electrochemical properties of the electrodes. CV was performed in the range of -0.2-0.8 V at a scan rate of 0.1 V·s^-1 in 0.1 mol·L^-1 KCl containing 1 mmol·L^-1 [Fe(CN)₆]^3- and [Fe(CN)₆]^4-. EIS was carried out with an Autolab electrochemical analyzer (Eco Chemie, Netherlands) in 20 mmol·L^-1 K₃[Fe(CN)₆] and K₄[Fe(CN)₆] (0.1 mol·L^-1 KCl) as the supporting electrolyte at a bias potential of 0.18 V, within the frequency range of 0.001 to 100 kHz. Secondly, chronoamperometry was performed at a potential of -0.1 V for 50 s in 0.1 mol·L-1 phosphate buffer solution (PBS, pH=7.4) containing 0.1 mol·L^-1 NaCl to establish the calibration plots of glucose. The amplitude was 50 mV with a pulse width of 0.167 s, and a pulse period of 0.5 s.

  1.5   Detection of glucose in real samples

  To investigate the practicability of the proposed sensor under clinical conditions, human serum samples were used as real samples to conduct the electrochemical measurements. The human serum samples were diluted by 10 times with 0.1 mol·L^-1 PBS (pH=7.4) solution. For glucose recovery in serum samples, different amounts of glucose were spiked into the above-diluted serum samples. Chronoamperometry measurements were carried out to detect the concentration of glucose using Pt/Au/AN electrodes. The recovery rates were obtained by the ratio of the calculated concentration to the spiked concentration.

  2.   Results and discussion

  2.1   Characterizations of the modified electrode

  The surface morphology of Pt/Au/AN electrode was characterized by SEM, as shown in Fig. 1. For the bare AN electrode, a smooth tip surface with a tip diameter of around 5 µm can be observed (Fig. 1A, 1B). It had a stainless steel metallic luster. In addition, we calculated that the diameter of the modified AN was about 160 µm at a 1.5 mm position from the tip (Fig. S1, Supporting information). After the modification of gold nanomaterials, Au/AN electrode exhibited a golden appearance (Fig. 1A). A stone-like morphology with an average diameter of 300 nm was further observed (Fig. 1C, 1D). Subsequently, the surface of Au/AN electrode was coated with the bimetallic Pt/Au nanoparticles through the electrodeposition strategy. It can be observed that Pt/Au/AN electrode displayed a grey slush appearance (Fig. 1A). A cabbage-like morphology was uniformly and densely distributed on the surface of AN electrode (Fig. 1E, 1F), which effectively increases the surface area of the electrode and then provides abundant electro-active sites for glucose oxidation^[35]. These characterizations demonstrated that the functional needle electrode was successfully fabricated. AN is integrated as the base layer and provides a large surface for follow-up growth of Au and Pt, developing needle electrodes with a large surface-to-volume ratio and much actives sites for the following determinations.

  Figure 1

  

  Figure 1.  (A) Bright-field image of bare AN, Au/AN, and Pt/Au/AN electrode; SEM images of AN electrode (B), Au/AN electrode (C, D), Pt/Au/AN electrode (E, F)

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  EIS is an effective tool to study the conductivity of modified electrodes as well as to characterize the modification process^[36]. The EIS spectrum contains a semicircle part and a linear portion, where the semicircle portion at high frequency represents the electron transfer limited process, and the linear part at low frequency represents a diffusion-limited process. According to the EIS data of the electrode, Randle′s equivalent circuit of the electrode can be obtained, which is consisted of ohmic resistance (R_s), double layer capacitance (CPE), and transfer resistance (R_ct), and Warburg impedance (Z_W). Among these components, the s_ct was calculated by the semicircle diameter of EIS spectrum, which controls the electron-transfer kinetics of the active species on the electrode surface. Fig. 2A and 2B show the typical Nyquist plots of AN, Pt/AN, Au/AN, and Pt/Au/AN electrodes, which were recorded in 0.1 mol·L^-1 KCl solution containing 20 mmol·L^-1 [Fe(CN)₆]^3- and [Fe(CN)₆]^4- at a frequency from 0.001 to 100 kHz at a potential of 0.005 V. As shown in Fig. 2A, the Nyquist plot of bare AN possessed a large semicircle portion that corresponded to a large electron transfer resistance (30 656.0 Ω), which means the electron transfer is difficult. After modification of Au or Pt, the electron transfer resistance was found to be 7.4 and 296.9 Ω, as shown in Fig. 2B. The downward trend of electron transfer resistance was ascribed to the excellent conductivity of Au or Pt, which could promote the electron transfer between solution and electrode interface. After further modification of Pt on Au/AN, the electron transfer resistance was reduced continuously and the value of electron transfer resistance was 3.2 Ω (Fig. 2B). The lowest electron transfer resistance value indicates the highest conductivity. The results indicate that the bimetallic Pt/Au nanoparticles are successfully modified onto the ANs and can effectively enhance the electrontransfer ability of electrodes compared with single metal. In addition, the Randle′s equivalent circuit of Pt/Au/AN electrode is shown in Fig.S2.

  Figure 2

  

  Figure 2.  EIS spectra of (A) bare AN and (B) different modified electrodes of Au/AN, Pt/AN, and Pt/Au/AN

  Inset in 2B: enlarged view of the modified electrodes at 0-100 Ω

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  CV is a useful technique for studying the electrochemical performance of electrode surfaces^[37]. Herein, the performance of four different electrodes, including AN, Pt/AN, Au/AN, and Pt/Au/AN were investigated by performing CV experiments (Fig. 3). For bare AN, it can be seen that there was no obvious redox peak, indicating the poor conductivity and electron transfer rate of [Fe(CN)₆]^3- and [Fe(CN)₆]^4- on AN. For Au/AN and Pt/AN, it was found that there were a pair of symmetrical redox peaks with a peak potential separation (ΔE_p) of 90 and 138 mV respectively, which could be attributed to Au and Pt nanoparticles for improving the electron transfer and enhancing the conductivity of the electrode. Moreover, Pt/Au/AN electrode generated a pair of symmetrical redox peaks with a peak potential separation (ΔE_p) of 90 mV. The peak current of Pt/Au/AN electrode increased by 4 times and 1.8 times compared to Pt/AN and Au/AN. These observations indicated that the bimetallic Pt/Au can increase the electron transfer rate more effectively than single metal, which may be due to the synergistic effect of bimetal. To confirm the best electrochemical performance of Pt/Au/AN electrode, the electrochemically active surface areas of different electrodes were calculated according to the Randles-Sevcik equation (Eq.1).

  $ {i_{\rm{p}}} = 2.69 \times {10^5}{n^{3/2}}Ac{D^{1/2}}{v^{1/2}} $

  (1)

  Figure 3

  

  Figure 3.  CV responses of bare AN, Au/AN, Pt/AN, and Pt/Au/AN electrodes at the scan rate of 50 mV·s^-1

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  Herein, i_p, n, A, c, D, and v represent the peak current, the number of electrons involved in the redox couple, electrode area, analyte concentration, analyte diffusivity, and scan rate, respectively. By calculation, the active areas of Pt/AN, Au/AN, and Pt/Au/AN were 0.014 5, 0.032 8, and 0.058 4 cm², respectively. It is generally known that the larger the electrode surface area, the better the performance. The largest surface area achieved at Pt/Au/AN verified the best electrochemical performance of Pt/Au/AN electrode compared with other electrodes.

  2.2   Electrooxidation of glucose on the modified electrodes

  Before implementing Pt/Au/AN electrode for glucose sensing, its electrochemical behavior was investigated by performing CV experiments in 0.1 mol·L^-1 PBS (pH=7.4) solution containing 0.1 mol·L^-1 NaCl in the presence and absence of glucose (5 mmol·L^-1 glucose of Fig. 4A; 5 and 10 mmol·L^-1 glucose of Fig. 4B) at a scan rate of 50 mV·s^-1 within the scan range from -0.65 to 0.5 V. No obvious redox peak was observed at bare AN and Pt/AN in the absence and presence of glucose, which suggests that these electrodes do not catalyze the oxidation of glucose in this potential window. In contrast, Au/AN generated two tiny oxidation peaks of glucose at -0.04 and 0.24 V, indicating low electrocatalytic activity of Au/AN electrode towards glucose oxidation. On the other hand, the CV of Pt/Au/AN without glucose has three simple and clear parts: two pairs of redox peaks at negative potentials are ascribed to hydrogen adsorption/desorption peaks, a flat double layer region at intermediate potentials, and metal oxide formation/reduction peaks at positive potentials (Fig. 4B). After the addition of glucose, the CV curves of Pt/Au/AN were similar at various concentrations of glucose, indicating that Pt/Au/AN electrode is stable for glucose electro-oxidation under the CV method. Meanwhile, Pt/Au/AN electrode had multiple obvious oxidation peaks in the forward scan, among which three obvious oxidation peaks were located at -0.46, -0.37, and -0.09 V. According to the number of oxidation peaks, it can be inferred that the oxidation of glucose may be completed in multiple steps. The first oxidation peak may be caused by the adsorption and dehydrogenation of glucose on the electrode, and the following oxidation peaks may be attributed to the further oxidation of intermediate products^[38]. The reduction peak at 0.09 V in the backward scan may be caused by the reduction of surface oxides at high potential. As the surface oxide decreased, more active sites were exposed to the oxidation of glucose, resulting in a larger oxidation current. Therefore, according to the electrocatalytic results detected by CV, the prepared Pt/Au/AN electrode provided a higher catalytic activity toward glucose oxidation than AN, Pt/AN, and Au/AN.

  Figure 4

  

  Figure 4.  CV responses of (A) bare AN, Au/AN, Pt/AN, and (B) Pt/Au/AN in 0.1 mol·L^-1 PBS (pH=7.4) solution containing 0.1 mol·L^-1 NaCl in the presence and absence of glucose at the scan speed of 50 mV·s^-1; (C) CV response of Pt/Au/AN electrode at different scan rates from 10 to 1 000 mV·s^-1 in 0.1 mol·L^-1 PBS (pH=7.4) containing 0.1 mol·L^-1 NaCl and 5 mmol·L^-1 glucose; (D) Plot of i_p at -0.09 V vs square root of scan rate

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  Furthermore, the glucose oxidation process on Pt/Au/AN electrode was also discussed, as shown in Scheme 1C. Firstly, glucose was oxidized to gluconolactone. Then, gluconolactone was directly oxidized to gluconic acid^[39]. To further investigate the mechanism, the effect of scan rate on oxidation peak current at Pt/Au/AN electrode was measured in a range of 10 to 1 000 mV·s^-1, in which 0.1 mol·L^-1 PBS (pH=7.4) solution containing 0.1 mmol·L^-1 NaCl and 5 mmol·L^-1 glucose (Fig. 4C). It can be seen that the anodic peak currents increased with the increase of the scanning rate. As shown in Fig. 4D, the anodic i_p at -0.09 V showed a good linear relationship with the square roots of scan rates (v). The linear regression equation is i_p=62.65v^1/2-1.41 (R²=0.998), implies that the electrocatalytic for glucose oxidation on Pt/Au/AN electrode is a diffusioncontrolled electrochemical process.

  2.3   Optimization of the experimental conditions

  According to the above discussion, Pt/Au/AN electrode exhibited great electrocatalytic performance toward glucose oxidation. To further improve the electrochemical performance of Pt/Au/AN electrode for detecting glucose, several experimental parameters, including electrodeposition potential, time, the concentration of H₂SO₄, the molar ratio of HAuCl₄ to H₂PtCl₆ were optimized. CV was carried out and the current difference (Δi_p) was recorded at Pt/Au/AN electrode in the absence and presence of 5 mmol·L^-1 glucose. Firstly, the potential and time for the electrodeposition procedure were optimized. As shown in Fig. 5, the value of Δi_p increased significantly with increasing the electrodeposition potential and time. The reason may be attributed to the increase in the amount of metal nanoparticles deposited onto the electrode surface to improve the surface area and conductivity of the electrode. When the deposition potential and time increased to a certain extent, the value of Δi_p began to decrease, which may be caused by the accumulation of metal covering the active sites. Therefore, the best potential and time of the electrodeposition procedure for Au were -8.0 V and 180 s, and the best electrodeposition parameters of Pt were -0.8 V and 300 s.

  Figure 5

  

  Figure 5.  Effect of (A) electrodeposition potential for Au, (B) electrodeposition potential for Pt, (C) electrodeposition time for Au, and (D) electrodeposition time for Pt on the value of Δi_p

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  Subsequently, the effect of H₂SO₄ concentration on the electrochemical catalysis of Pt/Au/AN electrode toward glucose was studied (Fig. 6A and 6B). It was found that the current elevated with increasing H₂SO₄ concentration, and started to level off after a certain value. Based on the experimental results, the optimized H₂SO₄ concentrations for the electrodeposition of Au and Pt were 0.2 and 0.4 mol·L^-1. Most importantly, the influence of the molar ratio of HAuCl₄ to H₂PtCl₆ on the electrochemical catalysis of Pt/Au/AN electrode toward glucose was explored. The total concentration of HAuCl₄ and H₂PtCl₆ was controlled to be 2 mmol·L^-1. As shown in Fig. 6C, the value of Δi_p increased with decreasing the molar ratio of H₂PtCl₆ to HAuCl₄, which may be due to the large increase in the poisoning ability of Pt catalyst resulting from the increased loading of Au. After reaching the maximum value of Δi_p at the molar ratio of 3:5 (H₂PtCl₆ to HAuCl₄), the electrocatalytic activity of Pt/Au/AN electrode started falling, which implies that the electrocatalytic activity may be mainly determined by the amount of Pt. The results indicated that the contact interface of the bimetallic Pt/Au is the vital electrocatalytic site for glucose oxidation.

  Figure 6

  

  Figure 6.  Effects of the concentration of H₂SO₄ in preparing HAuCl₄ solution (A), and H₂PtCl₆ solution (B), the concentration ratio of H₂PtCl₆ to HAuCl₄ (C) on the value of Δi_p

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  2.4   Amperometric detections of glucose on Pt/Au/AN electrode

  To valid the potential application of Pt/Au/AN electrode, the capacity of Pt/Au/AN electrode for the determination of different glucose concentrations was measured by chronoamperometry at a potential of -0.1 V for 50 s in 0.1 mol·L^-1 PBS (pH=7.4) solution containing 0.1 mol·L^-1 NaCl. As shown in Fig. 7A, the amperometric current of Pt/Au/AN increased linearly with the increase of glucose concentration (c) from 0.1 to 35 mmol·L^-1. The linear calibration curve of Pt/Au/AN can be expressed as i=0.968c^1/2+0.133 (R²=0.996) and the limit of detection (LOD, S/N=3) was calculated to be 76.3 µmmol·L^-1. The reason why Pt/Au/AN electrode possesses high sensitivity and wide linear range may be attributed to the synergy of bimetals and their contact interface. Meanwhile, the comparison between the previous electrochemical sensors for non-enzyme determination of glucose is shown in Table 1. Our proposed biosensor exhibited an acceptable detection limit and a wider detection range, which helps diagnose hypo and hyperglycemic cases with highly significant in the real-time monitoring of glucose. Moreover, the preparation of Pt/Au/AN electrode is great cheap, and simple, which is expected to be used for actual sample analysis.

  Figure 7

  

  Figure 7.  (A) Amperometric response of Pt/Au/AN electrode to different glucose concentrations (0, 0.1, 0.2, 0.5, 1, 2, 4, 6, 8, 10, 15, 20, 25, 30, 35 mmol·L^-1) at 0.1 V; (B) Calibration curves of current vs square root of glucose concentration from Pt/Au/AN electrode by chronoamperometry

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  Table 1

  

  Table 1.  Comparison of non-enzyme glucose sensors with different materials

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  Material	Linear range/(mmol·L-1)	LOD/(µmol·L-1)	Sensitivity/(µA·mmol-1·L·cm-2)	Ref.
  Pt/Au nano-alloy/Pt	1.39-13.9	482	2.82	[39]
  Pt/Au/BDD	0.01-7.5	6.5	—	[24]
  PtAu/GCE	0.01-10	3	—	[16]
  Cu-Al LDH/SPE	3-27	17.6	1 111.15	[13]
  Cu2O-Au/Cu foil	1-20	10	2 430	[35]
  Pt/Au/AN	0.1-35	76.3	16.58	This work

  2.5   Selectivity, reproducibility, and stability of Pt/Au/AN electrode

  Selectivity, which is a crucial indicator for sensors, can evaluate the ability to distinguish the interference and other targets. For the detection of glucose in human serum, the main interfering substances are some electrically active substances, such as AA, UA, 4-acetaminophen (AP), and DA. To assess the selectivity of Pt/Au/AN electrode, interferences have been added into 0.1 mol·L^-1 PBS (pH=7.4) solution containing 0.1 mol·L^-1 NaCl and 5 mmol·L^-1 glucose, and then the current intensity changes were recorded. As illustrated in Fig. 8, it was found that an obvious current response was observed after injection of glucose, while the current response did not change obviously after adding 0.1 mmol·L^-1 AA, 0.5 mmol·L^-1 UA, 0.2 mmol·L^-1 AP, and 0.2 mmol·L^-1 DA respectively. Therefore, it shows that the sensor has high selectivity and anti-interference from the anti-interference experiments.

  Figure 8

  

  Figure 8.  Amperometric response of Pt/Au/AN electrode upon the injection of glucose, AA, UA, AP, DA, and glucose

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  Reproducibility and stability are also important evaluation indicators for sensors. After comparing the current response of six Pt/Au/AN electrodes in 0.1 mol·L^-1 PBS (pH=7.4) solution containing 0.1 mol·L^-1 NaCl and 5 mmol·L^-1 glucose by CV, the result of relative standard deviation (RSD) from six parallel experiments was 4.02%, which shows good repeatability. For the evaluation of stability, the prepared Pt/Au/AN electrode was stored at room temperature for 15 d to compare the response current before and after 15 d. The result showed that Pt/Au/AN electrode remained at 90.5% of the initial current, indicating that the electrode has great stability.

  2.6   Determination of glucose in real sample

  Pt/Au/AN sensor was used to test human serum samples to further verify its potential application. Firstly, each real sample was diluted by 10 times with PBS (pH=7.4) and then the concentrations of glucose were measured by chronoamperometry. The obtained results were compared with the concentration value measured by Hangzhou Cancer Hospital. Finally, glucose was added to the diluted sample and the concentration of glucose was measured by chronoamperometry again. The measurement results are shown in Table 2. It can be seen that RSD and recovery were acceptable, indicating that the sensor has good accuracy for glucose detection and a wide range of potential applications in clinical monitoring.

  Table 2

  

  Table 2.  Using Pt/Au/AN sensor to measure glucose in real human serum samples

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  Sample	Hospital measured concentration/(mmol·L-1)	Detected concentration/(mmol·L-1)	RSD/% (n=6)	Added concentration/(mmol·L-1)	Detected concentration/(mmol·L-1)	Recovery/%	RSD/% (n=6)
  1	5.07	4.96	5.42	5	9.84	97.6	4.76
  2	4.66	4.43	8.63	5	9.70	105.4	5.76
  3	4.70	4.75	1.59	5	9.87	102.4	2.17
  4	4.74	4.59	4.65	10	13.64	90.5	4.84
  5	4.99	5.16	3.92	10	15.77	106.10	5.27
  6	4.36	4.50	2.66	10	15.29	107.90	8.45

  3.   Conclusions

  In this work, we successfully modified bimetallic Pt/Au cabbage - like nanoparticles onto acupuncture needles by electrodeposition, which can be used to construct a non - enzyme glucose sensor (Pt/Au/AN). Compared with Pt/AN or Au/AN sensors, Pt/Au/AN sensor has better electrocatalytic performance for glucose oxidation, high sensitivity, and selectivity, which is mainly due to the synergistic effect and their contact interface of Pt and Au. Under the optimized conditions, Pt/Au/AN electrode has a good linear relationship over the glucose concentration ranging from 0.1 to 35 mmol·L^-1 with the detection limit of 0.076 3 mmol·L^-1. Finally, the obtained sensor exhibited an excellent ability to detect glucose in human serum samples. In addition, Pt/Au/AN sensor was constructed based on the acupuncture needle, which possesses a low cost and simple preparation method. In general, these features make Pt/Au/AN sensor have a wide range of potential applications in clinical monitoring.

  Supporting information is available at http://www.wjhxxb.cn

  
  Acknowledgments: This work is supported by the Science and Technology Program of Zhejiang Province of China (Grants No. LGF22H200012, LGC20H200001, LGF22B050008), the Medical and Health Technology Development Program of Zhejiang Province (Grant No. 2020KY727), the Program for Science and Technology of Jiaxing (Grant No. 2017AY33086). We express our heartfelt thanks to them for their financial support. There is no conflict of interest, financial or otherwise. ^#共同第一作者。
  
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  Scheme 1  (A) Fabrication procedure of Pt/Au/AN electrode; (B, C) Mechanism of glucose oxidation on Pt/Au/AN electrode

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  Figure 1  (A) Bright-field image of bare AN, Au/AN, and Pt/Au/AN electrode; SEM images of AN electrode (B), Au/AN electrode (C, D), Pt/Au/AN electrode (E, F)

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  Figure 2  EIS spectra of (A) bare AN and (B) different modified electrodes of Au/AN, Pt/AN, and Pt/Au/AN

  Inset in 2B: enlarged view of the modified electrodes at 0-100 Ω

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  Figure 3  CV responses of bare AN, Au/AN, Pt/AN, and Pt/Au/AN electrodes at the scan rate of 50 mV·s^-1

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  Figure 4  CV responses of (A) bare AN, Au/AN, Pt/AN, and (B) Pt/Au/AN in 0.1 mol·L^-1 PBS (pH=7.4) solution containing 0.1 mol·L^-1 NaCl in the presence and absence of glucose at the scan speed of 50 mV·s^-1; (C) CV response of Pt/Au/AN electrode at different scan rates from 10 to 1 000 mV·s^-1 in 0.1 mol·L^-1 PBS (pH=7.4) containing 0.1 mol·L^-1 NaCl and 5 mmol·L^-1 glucose; (D) Plot of i_p at -0.09 V vs square root of scan rate

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  Figure 5  Effect of (A) electrodeposition potential for Au, (B) electrodeposition potential for Pt, (C) electrodeposition time for Au, and (D) electrodeposition time for Pt on the value of Δi_p

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  Figure 6  Effects of the concentration of H₂SO₄ in preparing HAuCl₄ solution (A), and H₂PtCl₆ solution (B), the concentration ratio of H₂PtCl₆ to HAuCl₄ (C) on the value of Δi_p

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  Figure 7  (A) Amperometric response of Pt/Au/AN electrode to different glucose concentrations (0, 0.1, 0.2, 0.5, 1, 2, 4, 6, 8, 10, 15, 20, 25, 30, 35 mmol·L^-1) at 0.1 V; (B) Calibration curves of current vs square root of glucose concentration from Pt/Au/AN electrode by chronoamperometry

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  Figure 8  Amperometric response of Pt/Au/AN electrode upon the injection of glucose, AA, UA, AP, DA, and glucose

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  Table 1.  Comparison of non-enzyme glucose sensors with different materials

  Material	Linear range/(mmol·L-1)	LOD/(µmol·L-1)	Sensitivity/(µA·mmol-1·L·cm-2)	Ref.
  Pt/Au nano-alloy/Pt	1.39-13.9	482	2.82	[39]
  Pt/Au/BDD	0.01-7.5	6.5	—	[24]
  PtAu/GCE	0.01-10	3	—	[16]
  Cu-Al LDH/SPE	3-27	17.6	1 111.15	[13]
  Cu2O-Au/Cu foil	1-20	10	2 430	[35]
  Pt/Au/AN	0.1-35	76.3	16.58	This work

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  Table 2.  Using Pt/Au/AN sensor to measure glucose in real human serum samples

  Sample	Hospital measured concentration/(mmol·L-1)	Detected concentration/(mmol·L-1)	RSD/% (n=6)	Added concentration/(mmol·L-1)	Detected concentration/(mmol·L-1)	Recovery/%	RSD/% (n=6)
  1	5.07	4.96	5.42	5	9.84	97.6	4.76
  2	4.66	4.43	8.63	5	9.70	105.4	5.76
  3	4.70	4.75	1.59	5	9.87	102.4	2.17
  4	4.74	4.59	4.65	10	13.64	90.5	4.84
  5	4.99	5.16	3.92	10	15.77	106.10	5.27
  6	4.36	4.50	2.66	10	15.29	107.90	8.45

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