Introduction

Lead has severe effects on human health and the environment [1,2,3]. Hence, it is important to develop highly selective, sensitive and cost-effective lead(II) assays. A number of analytical techniques such as fluorescent, electrochemical and colorimetric ones has been developed for lead(II) [4,5,6,7]. Among them, fluorometric methods, including both DNAzyme and aptamer-based approaches, are ideal due to their distinct advantages such as simple, sensitive and inexpensive, and can determine concentrations to trace levels [7,8,9]. In this type of assays, various fluorescent probes like organic dyes or fluorophores have been employed for the detection of lead(II) [3, 7, 10, 11]. However, organic dyes or fluorophores might suffer from small Stokes shift, short fluorescence lifetime and emission wavelength, and poor photostability. These resulted in certain limitations including low sensitivity and poor selectivity, and so on.

Metal nanoclusters have drawn considerable attention due to their molecular-like fluorescence emission with size-dependent spectra, large Stokes shift, good photostability and high quantum efficiency [12, 13]. These features make them as the ideal fluorescent probes for wide applications in the biomedical and environmental fields [14,15,16]. Also, their low-cost and easy synthesis make them an attractive alternative to common fluorophores [17, 18]. In particular, gold nanoclusters (AuNCs) as a promising class of fluorescent probes have been employed in chemical and biological detection [15, 18, 19]. Ongoing efforts have been devoted to the facile synthesis of AuNCs using various methods with the increasing usage of biomolecules, such as protein, enzyme and oligonucleotide, as the environmentally benign templates [12, 15, 17]. Nevertheless, no reports were found employing protamines as template for synthesizing AuNCs for detecting metal ions. Furthermore, some of nanoclusters-based fluorescent probes possess relatively low quantum efficiency and poor photostability. Therefore, the development of new fluorescent nanomaterials with high quantum efficiency and photostability remains a major challenge.

Protamines are small arginine-rich basic proteins that condense the DNA in mature spermatozoa [20]. They are purified by the mature testes of certain fishes such as salmon and herring. Protamines are known to exhibit cell penetrating activity [21], and utilized for efficient delivery of DNA in transfection studies of cells [22]. Moreover, the attachment of DNA to gold microparticles by protamines also enhances the resistance to temperature and protease degradation [23]. In addition, protamines are an environmentally-benign reducing and stabilizing reagent, and can offer great water solubility and natural biocompatibility. Their 3D complex structure can also be easily conjugated with other systems. These characteristics of protamines can be ideally used as template for the preparation of metal nanoclusters.

In this article, we utilized protamine as template for synthesizing gold nanoclusters (PRT-AuNCs) that possess high quantum efficiency and photostability, and large Stokes shift. Taking advantage of the prepared PRT-AuNCs as fluorescence nanoprobe, we found that Pb2+ ions can selectively and sensitively quench their fluorescence by Pb2+ –Au+ interaction. Thereby, an inexpensive, selective and sensitive approach for lead(II) detection was established. This strategy is not only helpful to develop novel fluorescence nanoprobes, but also irradiative to discover new applications of AuNCs in environmental analysis and biomedical field.

Materials and methods

Reagents

Protamine sulfate, HAuCl4·4H2O and sodium citrate were purchased from Sinopharm Chemical Reagent Co., Ltd. (Shanghai, China, http://www.instrument. com.cn/). Lead(II) standard was bought from Beijing century Bioko Biological Technology Co., Ltd. (Beijing, China, http://www.bzwz.com/) and the concentration of its stock solution is 482.6 μM. 4-(2-hydroxyl- ethyl)piperazine-1-ethanesulfonic acid sodium salt (HEPES) and Tris(hydroxylmethyl)aminomethane (Tris) were obtained from Aladdin Chemistry Co., Ltd. (Shanghai, China, http:// www.aladdin-reagent.com/). Sodium citrate-hydrochloric acid buffer (10 mM, pH 6.5) was employed to control the acidity of the solution. All chemicals used were analytical grade, and doubly distilled water (18.2 MΩ cm) was utilized throughout this work.

Apparatus

All the fluorescence spectra were acquired on a Hitachi F-4500 spectrofluorometer (Tokyo, Japan) equipped with a quartz cuvette (10 mm). The slit widths of both excitation and emission were kept at 5 nm and 10 nm, respectively. Transmission electron microscopy (TEM) images were obtained by a FEI Tecnai G2 F20 S-TWIN Field emission transmission electron microscope (Oregon, USA). X-ray photoelectron spectroscopy (XPS) was measured with a Thermo Scientific Escalab 250Xi X-ray photoelectron spectrometer (Waltham, UK). A pH meter (Sartorius AG, Germany) was utilized for pH adjustment.

Synthesis of the protamine-gold nanoclusters

Protamine-gold nanoclusters (PRT-AuNCs) were synthesized similarly by a one-step synthesis procedure. In brief, an aqueous solution of 25 mL of 2.5 mg mL−1 protamine sulfate was added to an equal volume of 10 mM HAuCl4. The mixture was vigorously stirred for 2 min at 37 °C, followed by adding 2.5 mL of 1 M NaOH. The resulting solution was incubated for 12 h at 37 °C under mild stirring. A yellowish brown solution appeared in the reaction flask, prompting the formation of the yellowish green fluorescent PRT-AuNCs. The solution was then dialyzed in water for 24 h, and the aggregation occured, which led to a decreased fluorescence intensity. So, the PRT-AuNC solution was directly employed in the experiments without further purification.

X-ray photoelectron spectroscopy measurement

X-ray photoelectron spectroscopy (XPS) measurements were carried out on a Thermo Scientific Escalab 250Xi X-ray photoelectron spectrometer (Waltham, UK) with a monochromated Al K alpha X-ray source (1468.6ev). The solutions of PRT-AuNCs and PRT-AuNCs-Pb(II) were prepared respectively. The residues obtained by centrifuging were coated onto the face of prepared 1 × 1 cm glass slides, then soaked in 10% nitric acid overnight, and diped in the 30% H2O2 for another 8 h. Thereafter, the samples were taken into the vacuum drying oven for drying before measurement.

Fourier-Transform Infrared (FTIR) spectrum measurement

FTIR spectra were measured by Shimadzu IRPrestige-21 FTIR spectrometer (Kyoto, Japan), equipped with a KBr beam splitter, a standard source and a DTGS Peltier-cooled detector. The solutions of PRT, PRT-AuNCs and PRT-AuNCs-Pb(II) were prepared respectively. Thereafter, the resulting solutions were drawed onto the middle of two KBr pellet with capillary tube. The KBr pellet spectra of samples were obtained in the range from 4000 to 400 cm−1 with a resolution of 4 cm−1.

General procedures

Into a 2-mL Eppendorf-type plastic tube, a 40 μL PRT-AuNC solution and an appropriate amount of lead(II) solution were added into a 15 μL citrate-HCl buffer of pH 6.5, followed by adding water to a volume of 500 μL. After mixing thoroughly and incubating for 20 min at 35 °C, the mixture was cooled to the room temperature. Subsequently, fluorescence spectra were measured by scanning in the wavelength range from 550 to 650 nm at λex = 300 nm, and the spectral bandwidths of both the excitation and emission slits were set to 5 nm and 10 nm, respectively. The fluorescence intensities were recorded at λmax 599 nm, representing as ΔF = F0 – F, here F0 and F were the fluorescence intensities of the system without and with lead(II), respectively.

Results and discussion

Choice of materials

The selection of an appropriate fluorescent nanoprobe for fluorescence detection of lead(II) is particularly important. In this work, we successfully synthesized PRT-AuNCs by one-pot method utilizing protamine as both a stabilizer and a reducing agent, which were employed to function as a fluorescent nanoprobe for lead(II). The average diameter of the PRT-AuNCs estimated from transmission electron microscopy (TEM) is d = 1.30 ± 0.20 nm for n = 70 (Fig. S1, see Electronic Supplementary Material, ESM). The aqueous solutions exhibit yellowish green fluorescence under 365 nm ultraviolet light (Fig. 1a). The absorption spectrum of the PRT-AuNCs shows no characteristic surface plasmon absorption of gold nanoparticles (data not shown), and their fluorescence band is centered at 599 nm upon excitation at 300 nm (Fig. S2, ESM). This Stokes shift value is about 1.5–2.5 times larger than that of previously reported GSH-AuNCs [24], GSH-AgNCs [25] and BSA-AuNCs [26]. The large Stokes shift can prevent spectral crosstalk, which therefore enhances the detection signal. Furthermore, the fluorescence intensity of the PRT-AuNCs solution as nanoprobe remained almost constant even one month after preparation, demonstrating that they are fairly stable compared to other AuNPs- and AuNCs-based nanomaterials (Fig. S3, ESM). These features of the PRT-AuNCs including molecular-like fluorescence emission, large Stokes shift, good photostability and high quantum efficiency make them an attractive alternative to common fluorophores or other fluorescent metal nanoclusters. Thereby, the PRT-AuNCs were chosen as fluorescence nanoprobe in this experiment. Additionally, the effect of the concentration of the PRT-AuNCs on the Pb(II) assay was also investigated. Figure 1b shows that ΔF values enhanced promptly with the raise of the PRT-AuNCs concentrations from 10 to 40 μL, reaching a maximum upon addition of 40 μL PRT-AuNCs solution. Thus, an appropriate amount of the PRT-AuNCs is requisite for the fluorescence assay of Pb(II). However, when the PRT-AuNCs concentrations were higher than the optimum one, the ∆F values of the detection system decreased gradually. Hence, a 40 μL solution was chosen as the optimum assay condition.

Fig. 1
figure 1

Visual observation of PRT-AuNCs (a), effects of PRT-AuNC concentration (b), buffer solutions (c) and pH (d) on Pb(II) assay, fluorescence spectra of the PRT-AuNCs-Pb(II) system (e), and Stern–Volmer plot at three different temperatures (f). a 365 nm UV light, cPb(II) = 1.5 μM, VPRT-AuNCs = 50 μL; b cPb(II) = 1.5 μM; c cPb(II) = 1.5 μM, cHEPES = cTris-HAc = 10.0 mM (pH 6.5), VPRT-AuNCs = 50 μL; d cPb(II) = 1.5 μM, VPRT-AuNCs = 50 μL. e λex = 300 nm, VPRT-AuNCs = 40 μL, cPb(II) (μM)/(a–h): 0.00, 0.10, 1.00, 2.00, 4.00, 8.00, 10.00, 12.00. f λex/λem = 300/599 nm, VPRT-AuNCs = 40 μL. (a), (b), (c), (e) and (f): ccitrate-HCl = 10.0 mM (pH 6.5). The error bars represent the standard deviations of three repetitive measurements

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The effects of the buffers citrate-HCl, HEPES and Tris-HAc were also investigated. The experimental result revealed that citrate-HCl was more favorable than others for the detection of Pb(II) ions (Fig. 1c). Therefore, citrate-HCl buffer was utilized in the experiments. Figure 1d displays that the ∆F went up with the increase of pH, and reached a maximum in the citrate-HCl buffer of pH 6.5. This result demonstrates that the interaction of the PRT-AuNCs with Pb2+ is highly pH dependent. The explanation may be that the hydrolyzed species such as Pb(OH)2 start to dominate at pH > 7.0. Thus, the citrate-HCl buffer of pH 6.5 was chosen for controlling the acidity of the solution.

Mechanism of the lead(II) induced fluorescence quenching

The analytical principle for this strategy is shown in Scheme 1. It was reported that some of diatomic ions like Pb(II) have strong closed-shell d10–s2 interactions with Au+. This aurophilic interaction causes Pb atoms/ions to become deposited on the particle [23]. Inspired by these reports, we presume that the active sites on the PRT-AuNCs surfaces are Au+. Pb(II) probably can bind to Au+ on the PRT-AuNC surfaces owing to its strong aurophilic interaction. As a result, the fluorescence of the PRT-AuNCs can be quenched by Pb(II). Thereby, a “turn off” fluorescence assay for Pb(II) was established by the PRT-AuNCs as a novel fluorescent nanoprobe.

Scheme 1
scheme 1

Schematic illustration of the fluorescence assay of lead(II) using PRT-AuNCs

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To prove above-mentioned supposition, a series of control experiments were performed. Firstly, the interaction of the PRT-AuNCs with Pb(II) was investigated by fluorescence spectra. Figure 1e shows that the maximum fluorescence for the PRT-AuNCs occurs at 599 nm (curve a). After Pb(II) was added to the solution, the signal of the system decreased gradually with increasing Pb(II) concentrations (curves b-h). The reason may be that the interaction of Pb2+ ions with the Au+ on the PRT-AuNCs surfaces might lead to Pb2+ ions being deposited on the surfaces of the PRT-AuNCs as a result of strong aurophilic interactions [19, 23].

In order to verify the quenching mechanism, the fluorescence quenching data were analyzed with the hypothesis that dynamic quenching occurred. For dynamic quenching, the decrease in intensity is described by the Stern-Volmer equation of F0/F = 1+ Ksv ∙cQ [27], where, F0 and F represent the fluorescence intensities in the absence and the presence of quencher, cQ and KSV are the molar concentration of quencher and the Stern–Volmer quenching constant, respectively. Figure 1f illustrates the Stern–Volmer plots of the quenching of PRT-AuNCs fluorescence by Pb2+ at different temperatures, and the corresponding KSV are shown in Table 1. The results indicate the KSV is inversely correlated with temperature, prompting that the probable quenching mechanism of fluorescence of PRT-AuNCs by Pb2+ is not initiated by dynamic quenching, but probably by static quenching resulting from the formation of a complex [27].

Table 1 Stern-Volmer quenching constant of PRT-AuNCs-Pb(II) system at different temperatures
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The aurophilic interactions of Pb2+ was also confirmed by a control experiment. Fig. S4 (ESM) displays a weak absorption signal at 416 nm in the solution of AuNPs-ABTS-H2O2 (curve a), prompting that AuNPs possess weak catalytic activity on the oxidation of ABTS by H2O2. Upon the addition of Pb2+ with various concentrations into the solution of AuNPs-ABTS-H2O2, an obviously enhanced absorbance is observed (curves b and c). This implies that the interactions of Pb2+ with Au+ on the surface of the AuNPs leads to the formation of Au-Pb complex that possesses the peroxidase-like activity.

The data from X-ray photoelectron spectroscopy further demonstrated our hypothesis. Figure 2a shows that the two distinct components for Au 4f7/2 appear at binding energies of 84.03 eV and 84.98 eV, which are able to be identified as Au(0) and Au(I), respectively [1, 28, 29]. The presence of Pb2+ caused the obvious change of the peak area and the peak position of Au0 and Au+ (Fig. 2b), indicating the existence of the interaction of Pb2+ ions with the Au+ on the PRT-AuNCs surface. Fig. 2c illustrated that the binding energy of 136.67 eV/141.53 eV and 138.79 eV/143.65 eV for Pb 4f7/2/Pb 4f5/2 verified that both Pb0 and Pb2+ existed on the surface of the PRT-AuNCs when they were incubated with Pb2+ [1, 28, 29]. These results imply that a Pb-Au alloy might be formed on the surface of the PRT-AuNCs by Pb2+ –Au+ interaction [1, 19]. We suppose that sodium citrate played an important role in assisting the formation of the Pb-Au alloys.

Fig. 2
figure 2

XPS curves of Au0 and Au+ in the absence (a) and presence (b) of Pb(II), and Pb0 and Pb2+ in the presence of PRT-AuNCs (c). The black dashed line represents the raw data and the red and blue dashed lines are the fitting result. The original bands are able to be spilt into a few new peaks according to the binding energy

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To further verify the formation of a Pb-Au alloy, a transmission electron microscope (TEM) was employed to characterize the structure of Pb-Au complex. Figs. S5(a) and (b) (ESM) illustrate the TEM images before and after the addition of Pb2+. The TEM images reveal that the AuNCs in the presence of 0.5 μM Pb2+ ions (Fig. S5(b), ESM) almost remained same sizes and shapes in comparison with that in the absence of Pb2+ ions (Fig. S5(a), ESM). This implies the formation of monolayer or submonolayer of Pb–Au alloy on the PRT-AuNCs surfaces [19]. This result is consistent with that obtained by XPS.

The interaction of Pb2+ with Au+ on the PRT-AuNCs surface was also confirmed by Fourier-Transform Infrared (FTIR) spectra. Figure 3a describes that a wide and strong peak occurs at 3423.6–3473.8 cm−1, which is attributed to the stretching vibrations of O−H/N−H for protamine [30]. The peaks at 1639.5 cm−1 (C=O stretching) and 1060.8 cm−1 (C-N stretching) correspond to amide I and aliphatic amine regions [31], and no peaks in both amide II and amide III regions were found. Compared to protamine, PRT-AuNCs displayed an apparent broadening and enhancing O − H/N − H stretching, and an increased peak of C=O stretching vibration, accompanied by the appearance of some new peaks located at 2358.9 cm−1, 2073.5 cm−1, 1788 cm−1 and 1031.9 cm−1 (Fig. 3b). These results suggest that the Au+ on the AuNCs surface might bind to the N atoms in amide I and aliphatic amine regions to form Au-N bond by coordinating interaction, or/and Au-O bond via the interaction of Au+ with O atoms in C=O of PRT. When Pb2+ was added in the solution of PRT-AuNCs, the obvious decrease of the percent transmittances for the principal peaks like O−H/N−H and C=O were observed along with the narrowing of peak width for O−H/N−H (Fig. 3c). This observation suggests that the interaction of Pb2+ with Au+ might cause the destruction of the Au-N and Au-O bonds, accompanied by the formation of a Au-Pb alloy. On the other hand, Pb2+ ions might react with PRT in PRT−AuNCs to form the Pb − protamine complex, promoting more Pb2+ being deposited onto AuNCs to lead to the formation of a Au-Pb alloy on their surfaces [19]. Moreover, the appearance of peaks at 650–679 cm−1 might be correlated with the combination of Au+ with metals like Pb2+.

Fig. 3
figure 3

FTIR spectra for PRT-AuNCs-Pb(II) detection system. a PRT, b PRT-AuNCs, c PRT- AuNCs + Pb(II)

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Optimization of method

Effect of reaction temperature

Fig. S6 (ESM) indicates that a maximum ΔF value was accomplished at 35 °C, and then decreased promptly from 45 °C to 65 °C. The reason might be that a higher temperature probably was unfavorable for forming Pb − protamine complex and a Au-Pb alloy on PRT-AuNCs surfaces. Therefore, 35 °C was employed for the further experiments.

Effect of reaction time

Fig. S7 (ESM) illustrates that a maximum and stable signal was realized from 20 to 35 min. Thus, the reaction time of 20 min was selected for the further experiments.

Selectivity

To evaluate the selectivity of PRT-AuNCs nanoprobe toward Pb2+ ions, the effects of potentially interfering species were investigated under the identical conditions. The allowable quantity of the interfering ones was defined as an error not more than ±5%. Fig. S8 (ESM) indicates that 1000 times of Mg2+, K+, Na+, 100 times of Zn2+, Cu2+, Ca2+, Mn2+, Cd2+, NH4+, 80 times of Ag+, 50 times of UO22+, Hg2+, 20 times of Al3+, and 10 times of Fe3+ do not interfere with the detection of Pb2+. This observation manifested that the Pb2+ ions are able to interact preferentially with PRT-AuNCs under the optimized experimental condition. Fig. S8 (ESM) illustrates that 100 times of Ca2+ do not interfere with detecting Pb2+. However, the millimolar Ca2+ in real samples can interfere with the detection of Pb2+. The interference of Ca2+ can be obviated via heating to boil for 10 min, then filtering using 0.22 μm filter membrane.

Sensitivity

To validate the sensitivity of the nanoprobe toward Pb2+ ions, the calibration plot was established under the optimum conditions. The linear relationship of the value of ΔF (F0–F) versus Pb2+ concentration was from 80 nM to 15 μM (Fig. 4). The equations of linear regression are ΔF = 25.43 + 51.97c(10−7 M) and ΔF = 1020.5 + 5.432c (10−7 M), ranging from 80 nM to 2 μM and 2 μM to 15 μM with the correlation coefficients of 0.9970 and 0.9906, respectively. The relative standard deviations for the 11 determinations of 0.10 μM, 4.0 μM and 15 μM of Pb2+ were 1.6%, 2.5% and 1.9%, respectively. The detection limit (LOD) was estimated to be 24 nM according to the formula cL = 3σ /m (σ represents the standard deviation of the 11 blank measurements, and m is the slope of the calibration plot). This result is well below the maximum level of lead(II) permitted by the U.S. EPA for drinking water (72 nM) [8, 9], suggesting that this strategy can potentially be employed for monitoring the level of Pb2+ in environmental samples.

Fig. 4
figure 4

Calibration plot of ΔF versus Pb(II) concentration. λex/λem = 300/599 nm, VPRT-AuNCs = 40 μL, ccitrate-HCl = 10.0 mM (pH 6.5). The inset shows the calibration plot of Pb(II) concentration ranging from 0.80 × 10−7 M to 20.0 × 10−7 M. The error bars represent the standard deviations of three repetitive measurements

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Application to water samples

To test the feasibility of the nanoprobe for Pb2+ assay, five water samples were collected from the Xiangjiang river, the pond in University of South China and tap water. All samples were filtrated, heating to boil for 10 min. Thereafter, the samples were cooled to room temperature, filtering by using 0.22 μm filter membrane for following use.

Firstly, the matrix effects were assessed by comparing the recoveries of spiked samples to those of equivalent concentration in standard solution by this method. Experimental results revealed the 101.2 ± 1.83%, 97.9 ± 2.29% and 100.1 ± 2.58 recoveries of Pb2+ from the samples spiked with standard solutions containing different concentrations of Pb2+ (Table 2). Compared to relative standard Pb2+ concentration, and the t-values at 95% confidence level do not exceed the theoretical value. This verifies that the recoveries are not statistically different, suggesting the negligible effect of matrix on the fluorescence quenching of PRT-AuNCs by Pb2+. The reason may be due to the low amount of sample volume compared to the total analysis volume (equal to a 1:10 sample dilution), and also attributes to the high selectivity of the method. Therefore, samples are able to be detected without further cleanup or dilution.

Table 2 Matrix effects, the validation of method and recovery
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Next, the validation of the method by analyzing of lead ions in a standard reference materials was performed. The concentrations of Pb2+ ions in spiked samples (the certificated values of 0.150, 1.500 and 15.00 μM provided by Beijing century Bioko Biological Technology Co., Ltd.) are 0.1518 ± 0.004, 1.4685 ± 0.045, and 15.015 ± 0.351 μM (n = 6), respectively (Table 2). The relative recoveries of added lead(II) ranged from 97.9 ± 2.29% to 101.2 ± 1.83%, suggesting good accuracy of the method for Pb2+ detection in the samples.

Moreover, the inter-day and intra-day precisions were tested at three varying levels of 0.1, 4.0, and 15.0 μM Pb2+, and each one was evaluated in triplicate within the same day and on three consecutive days. The RSDs were 3.67%, 3.79% and 4.81% for the intra-day analysis, and 6.12%, 5.01% and 5.47% for inter-day analysis (n = 6), respectively.

Finally, the method was used for the detection of Pb2+ concentrations in real samples. It is found that the Pb2+ concentrations in the five water sample are too low to be detected by this method. However, an obvious decrease in fluorescent signal was observed upon the addition of Pb2+ standard solution to samples, and the results were shown in Table S1 (ESM). The result indicates that the method is able to be used for analyzing Pb2+ in real samples.

Comparison with other methods

Compared with other methods for Pb2+ assay, the detection limit obtained by this work was 2–10 times lower than those of most of fluorescent and colorimetric assays [32,33,34,35,36]. It is also comparable to that of the colorimetric one using Gallic acid–GNPs [37], but higher than that obtained by catechin-Au NPs) [19], 4-MB/ S2O32−-Au NP and 2-ME/S2O32−-Au NP [38, 39]. Next, this strategy consists of only a nanoprobe, avoiding the dual label step in comparison with methods based on aptamer and DNAzyme. This is obviously more convenient and economical. Moreover, this approach provides a wider linear range and high selectivity that arises from the strong Pb2+ –Au+ interaction (Table 3). Furthermore, this strategy would possess a potential for analyzing biosamples such as blood/serum, which always display strong background. In addition, heating the water sample to boiling for obviating the interference of mM levels of Ca2+ in real samples may make the Pb detection complicated. The further improvement of the selectivity and sensitivity of this Pb2+ assay system is underway.

Table 3 Comparison of this strategy with other methods for the determination of Pb(II)
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Conclusions

In summary, we have established a straightforward one-pot approach for synthesizing high fluorescent PRT-AuNCs by protamine as both a reducing and a stabilizing reagent. To the best of our knowledge, this is the first example of the use of PRT-AuNCs as nanoprobe for the fluorescence detection of Pb2+ ions, which provided a number of distinctive advantages. First, in terms of the cost, this method is obviously cheaper due to simplifying experimental procedure, without costly instruments. Second, this approach provides a wider linear range, lower detection limit and high selectivity that arises from the strong Pb2+ –Au+ interaction. Third, the feasibility of the strategy was verified by detecting lead(II) in real water samples. We also demonstrated the mechanism of the fluorescence quenching by lead(II). Based on the high fluorescence and peroxidase-like activity of PRT-AuNCs, this strategy is speculated to be able to become a universal detection platform using PRT-AuNCs as a nanoprobe for various targets in the future.