GR-5 DNAzyme Research Articles

A Self-Assembled G-Quadruplex/Hemin DNAzyme-Driven DNA Walker Strategy for Sensitive and Rapid Detection of Lead Ions Based on Rolling Circle Amplification.

Herein, a sensitive biosensor is constructed based on a novel rolling circle amplification (RCA) for colorimetric quantification of lead ion (Pb2+). At the detection system, GR5 DNAzymes are modified on the surface of an immunomagnetic bead, and Pb2+ is captured by the aptamer, inducing the disintegration of the GR5 DNAzyme and the release of the DNA walker. After the introduction of the template DNA, T4 DNA ligase, and phi29 DNA polymerase, an RCA is initiated for the sensitivity improvement of this method. Moreover, a G4-hemin DNAzyme is formed as a colorimetric signal, owing to its peroxide-like activity to catalyze the TMB-H2O2 substrate. Under the optimized conditions, the limit of detection (LOD) of this fabricated biosensor could reach 3.3 pM for Pb2+ with a concentration in the range of 0.01-1000 nM. Furthermore, the results of real samples analysis demonstrate its satisfactory accuracy, implying its great potential in the rapid detection of heavy metals in the environment.

Ultralow background one-pot detection of Lead(II) using a non-enzymatic double-cycle system mediated by a hairpin-involved DNAzyme

A double-cycle system has been developed for specifically detecting trace amounts of Pb2+ by significantly decreasing the background signal. The detection involves two types of RNA cleavage reactions: one using a Pb2+-specific GR5 DNAzyme (PbDz) and the other utilizing a newly constructed 10–23 DNAzyme with two hairpins embedded in its catalytic center (hpDz). The ring-structured hpDz (c-hpDz) exhibits significantly lower activity compared to the circular 10–23 DNAzyme without hairpin structures, which plays a crucial role in reducing the background signal. When Pb2+ is present, PbDz cleaves c-hpDz to its active form, which then disconnects the molecular beacon to emit the fluorescent signal. The method allows for rapid and sensitive Pb2+ detection within 40 min for 10 fM of Pb2+ and even as short as 10 min for 100 nM of Pb2+. Additionally, visual detection is possible through the non-crosslinking assembly of Au nanoparticles. The entire process can be performed in one pot and even one step, making it highly versatile and suitable for a wide range of applications, including food safety testing and environmental monitoring.

A ratiometric fluorescence platform for lead ion detection via RNA cleavage-inhibited self-assembly of three-arm branched junction

Heavy metal pollution can pose a threat to food safety and human health, and accurate quantification of heavy metal ions is a vital requirement. Emerging DNA nanostructures-based biosensors offer attractive tools toward ultra-sensitive or rapid analysis of heavy metal ions. However, the problems including complex design, severe reaction conditions and undesirable reliability are inevitable obstacle in advancing their extension and application. Herein, a ratiometric fluorescent platform was established for monitoring lead ion (Pb2+) in food based on dual Förster resonance energy transfer (FRET) and RNA cleavage-inhibited self-assembly of three-arm branched junction (TBJ). GR-5 DNAzyme was employed for Pb2+ recognition, and enzyme-free amplification technique catalytic hairpin assembly (CHA) served to form FRET probes-carried TBJ. The substrate strand (S) of DNAzyme triggered the generation of CHA-TBJ, and Pb2+-responsive cleavage of S hindered the assembly of CHA-TBJ, causing opposite changes in the FRET states of FAM/BHQ1 and ROX/BHQ2 pairs. The fluorescence responses were recorded through synchronous fluorescence spectrometry to indicate Pb2+ concentration, allowing sensitive and reliable identification of Pb2+ in the linear range of 0.05–5 ng mL−1 with the detection limit of 0.03 ng mL−1. The Pb2+ detection can be achieved under conventional reaction conditions, simple mixing procedures and one-step measurement operation. The approach can afford excellent specificity for Pb2+ against competing metal ions, and can be applied to analyze Pb2+ in tea samples with satisfactory results. This facile fluorescence platform shows a capable method for Pb2+ detection, and provides new avenue in the development of ratiometric approaches and DNAzyme strategies for monitoring heavy metal pollution, facilitating the transformation of DNAzyme-based biosensors for food safety control.

A triple amplification strategy using GR-5 DNAzyme as a signal medium for ultrasensitive detection of trace Pb2+ based on CRISPR/Cas12a empowered electrochemical biosensor

Lead ions (Pb2+) are a well-known toxic heavy metal that poses a significant threat to human health. Therefore, the development of a simple and ultrasensitive technique for detecting Pb2+ is essential. With their trans-cleavage properties, the newly discovered CRISPR-V effectors have become a potential high-precision biometric tool. In this regard, a CRISPR/Cas12a-based electrochemical biosensor (E-CRISPR) has been developed, which is combined with the GR-5 DNAzyme that can specifically recognize Pb2+. In this strategy, the GR-5 DNAzyme acts as a signal-mediated intermediary, which can convert Pb2+ into nucleic acid signals, thereby becoming single-stranded DNA that triggers strand displacement amplification (SDA) reaction. This is coupled with following activated CRISPR/Cas12a cleavage of the electrochemical signal probe, enabling cooperative signal amplification for ultrasensitive Pb2+ detection. The proposed method has a detection limit as low as 0.02 pM. Therefore, we have developed an E-CRISPR detection platform with GR-5 DNAzyme as a signal medium (called SM-E-CRISPR biosensor). This provides a method for the CRISPR system to specifically detect non-nucleic substances by converting the signal using a medium.

Nanozyme-catalysed CRISPR-Cas12a system for the preamplification-free colorimetric detection of lead ion

CRISPR-based detection was often based on the target preamplification to realize the high sensitivity. Here, we prepared a CRISPR-Cas12a system for the colorimetric detection of lead ion (Pb2+) based on the assistance of DNAzyme and nanozyme instead of preamplification. The recognition between GR-5 DNAzyme and Pb2+ could trigger the CRISPR-Cas12a system. MnO2 nanozymes connected with magnetic beads through single stranded DNA were prepared as the colorimetric signal probes and catalyst of CRISPR-Cas12a system for the strong oxidase-like activity inducing the color change of 3,3′,5,5′-tetramethylbenzidine. The nanozyme-catalysed CRISPR-Cas12a system could be used to detect Pb2+ through the color change with high specificity and sensitivity. The linear range of this approach was 0.8 nM–2500 nM, with a limit of detection of 0.54 nM. This method was applied for the detection of the Pb2+ in food samples indicating good accuracy and anti-interference ability.

A novel design of DNA duplex containing programmable sensing sites for nanopore-based length-resolution reading and applications for Pb2+ and cfDNA analysis.

Glass nanopore is an ideal candidate for biosensors due to its unique advantages such as label-free analysis, single-molecule sensitivity, and easy operation. Previous studies have shown that glass nanopores can distinguish different lengths of double-stranded DNA (dsDNA) at the same time with the length-resolution ability. Based on this, we proposed a novel design of a dsDNA block containing a programmable sensing site inside, which can be programmed to respond to different target molecules and cleaved into two smaller DNA blocks. When programming the sensing site with different sequences, for example, programming it as the substrate of GR-5 DNAzyme and CRISPR-Cas12a system, the DNA block could realize Pb2+ and cfDNA detection with the length-resolution ability of the glass nanopore. This strategy achieved a Pb2+ detection range from 0.5 nM to 100 nM, with a detection limit of 0.4 nM, and a BRCA-1 detection range from 1 pM to 10 pM, with a detection limit of 1 pM. The programable sensing site is easy to design and has strong expandability, which gives full play to the advantages of glass nanopore in length-resolution ability for dsDNA, and is expected to become an optional design for biosensing strategy for the glass nanopore as a biosensing platform.

Csm6-DNAzyme Tandem Assay for One-Pot and Sensitive Analysis of Lead Pollution and Bioaccumulation in Mice.

Lead contamination in the environment tends to enter the food chain and further into the human body, causing serious health issues. Herein, we proposed a Csm6-DNAzyme tandem assay (termed cDNAzyme) using CRISPR/Cas III-A Csm6 and GR-5 DNAzyme, enabling one-pot and sensitive detection of lead contamination. We found that Pb2+-activated GR-5 DNAzyme produced cleaved substrates that can serve as the activator of Csm6, and the Csm6-DNAzyme tandem improved the sensitivity for detecting Pb2+ by 6.1 times compared to the original GR-5 DNAzyme. Due to the high specificity of DNAzyme, the cDNAzyme assay can discriminate Pb2+ from other bivalent and trivalent interfering ions and allowed precise detection of Pb2+ in water and food samples. Particularly, the assay can achieve one-step, mix-and-read detection of Pb2+ at room temperature. We used the cDNAzyme assay to investigate the accumulation of lead in mice, and found that lead accumulated at higher levels in the colon and kidney compared to the liver, and most of the lead was excreted. The cDNAzyme assay is promising to serve as analytical tools for lead-associated environmental and biosafety issues.

Engineering DNAzyme strategies for fluorescent detection of lead ions based on RNA cleavage-propelled signal amplification

Based on the high recognition ability and flexible programmability of GR5 DNAzyme, two fluorescent biosensors were engineered for amplified detection of Pb2+ via incorporating Ti3C2TX MXenes and embedding 2-aminopurine (2-AP), respectively. The quencher-required approach relied on the DNA affinity and fluorescence quenching ability of Ti3C2TX MXenes. Benefiting from the low background signal modulated by Ti3C2TX MXenes, the sensitive determination of Pb2+ was achieved in the linear range of 0.2–10 ng mL−1 with the limit of detection (LOD) of 0.05 ng mL−1. The quencher-free approach combined the fluorescent trait of 2-AP embedded in DNA structure, and the RNA cleavage-propelled digestion process of Exonuclease I (Exo I) for signal amplification, indicating the sensitive detection of Pb2+ with the LOD as low as 0.02 ng mL−1 in the linear range of 0.1–10 ng mL−1. Both DNAzyme assays exhibited simple procedures, favorable specificity, rapid analysis, and satisfactory application in standard reference materials (lead in drinking water) and spiked water samples. The two fluorescent biosensors established in this work would not only provide theoretic fundament for DNA adsorption of Ti3C2TX MXenes and the design of 2-AP-embedded DNAzyme assays, but also hold a great potential for on-site monitoring of lead pollution in water samples.

Platform Formed from ZIF-8 and DNAzyme: "Turn-On" Fluorescence Assay for Simple, High-Sensitivity, and High-Selectivity Detection of Pb2.

Lead contamination has posed a potential threat to the environment and food safety, arousing extensive concern. In this work, we fabricated a novel fluorescent sensing platform based on zeolitic imidazolate framework-8 (ZIF-8) and DNAzyme for monitoring Pb2+ in water and fish samples. ZIF-8 was proposed as a fluorescence quencher with the advantages of simple synthesis, low cost, and high quenching efficiency. The Pb2+-dependent GR5 DNAzyme containing the large ssDNA loop can be adsorbed onto ZIF-8 accompanied by fluorescence quenching. Upon binding with Pb2+, GR5 DNAzyme was activated and cleaved, leading to the release of FAM-labeled 5-base ssDNA, which restored the fluorescence. The "turn-on" assay can detect Pb2+ through the one-pot procedure in the range of 0.01-10.0 nM with a detection limit of 7.1 pM. The platform is promising for on-site monitoring of Pb2+ owing to the excellent performance of high sensitivity, low background, strong anti-interference ability, and simple operation.

Signal-on fluorescent strategy based on RNA cleavage-inhibited catalytic hairpin assembly and photo-induced electron transfer for Pb2+ detection

A signal-on fluorescence biosensing strategy was developed by integrating GR-5 DNAzyme, catalytic hairpin assembly (CHA), photo-induced electron transfer (PET) between G-quadruplex (G4)/hemin DNAzyme and fluorophore. An oligonucleotide (cGRE), complementary to the enzyme strand (GRE), was applied to facilitate the separation of substrate strand (GRS) from GR-5 DNAzyme. The separated GRS acted as a trigger to initiate the CHA process and the formation of G4/hemin DNAzyme, resulting in the fluorescence quenching via PET. The Pb2+-responsive cleavage of GRS led to the suppression of the occurrence of CHA and the generation of G4/hemin DNAzyme, yielding the enhancement of the fluorescence. This DNAzymes-CHA-PET Pb2+ assay can be carried out in a single tube, endowed with the obvious advantages of facile design, simple procedure and excellent specificity. The quantitative determination of Pb2+ was achieved in a broad range of 0.1–150 ng mL−1 with the detection limit of 0.03 ng mL−1. This strategy was successfully applied to the analysis of standard reference materials (drinking water), spiked water and fish samples. The ingenious design proposed herein provides a new method for routine screening of heavy metals, and also opens up new avenues for the development and application of PET biosensors based on G4/hemin DNAzyme.

Label-free electrochemical biosensor based on GR5 DNAzyme/Ti3C2Tx Mxenes for Pb2+ detection

• The interactions of Ti 3 C 2 T x Mxenes toward different DNA molecules were studied. • Pb 2+ could enhance the oxidation peak current of GR5 DNAzyme/Ti 3 C 2 T x Mxenes GCE. • Label-free sensor was fabricated based on GR5 DNAzyme/Ti 3 C 2 T x Mxenes GCE. • This sensor was successfully applied to detect Pb 2+ in rat and chicken samples. The interactions between nucleic acids and nanomaterials have been often utilized to design the electrochemical biosensor due to easy preparation, adsorptive tunability and high sensitivity. In this study, we have prepared a new electrochemical biosensor for Pb 2+ detection based on GR5 DNAzyme/Ti 3 C 2 T x Mxenes modified glassy carbon electrode (GR5 DNAzyme/Ti 3 C 2 T x Mxenes-GCE). GR5 DNAzyme is an RNA-cleavage-type DNAzyme that shows high specificity towards Pb 2+ . With the presence of Pb 2+ , the cleavage of GR5 DNAzyme is activated at a ribo -adenine (rA) site in the substrate stranded DNA, leading to the unwinding of double-helix structure. This can enhance the adsorptive property of Ti 3 C 2 T x Mxenes toward GR5 DNAzyme, which favors the ion intercalation into the Ti 3 C 2 T x Mxenes and induces the increase of oxidation peak current. Therefore, we have successfully applied GR5 DNAzyme/Ti 3 C 2 T x Mxenes-GCE to detect Pb 2+ with a linear range from 0.5 to 32 nM and a detection limit of 0.1 nM. This electrochemical biosensor is simple, sensitive and selective for Pb 2+ determination that it should find some potential applications in food safety inspection and environmental monitoring.

Lead-start isothermal polymerase amplification controlled by DNAzymatic switches.

As DNA polymerases are even active at ambient temperature, there is inevitable non-specific amplification; to avoid the undesired amplification of analytes, a heat activation-based polymerase chain reaction (PCR), called hot-start PCR, is widely used to be highly precise and quantitative in detection. Unlike thermocycling amplification, isothermal amplification, compatible for point-of-care (PoC) tests, cannot be benefited by the heat-activation technique, making the method qualitative rather than quantitative. In this work, we newly developed a lead ion (Pb2+) activation technique, called lead-start isothermal amplification, allowing on-demand activation or deactivation of DNA polymerases at room temperature. We systematically correlated the DNA polymerase inhibition by the TQ30 aptamer with Pb2+-responsive strand cleavage by the GR5 DNAzyme, and relying on the type of interconnectors, Pb2+ successfully served as an initiator or a terminator of isothermal DNA amplification. Our lead-start isothermal amplification was exceptionally Pb2+-specific, dramatically increasing the enzymatic activity of DNA polymerase (>25 times) only by Pb2+ introduction. Despite one-by-one sample preparation, a number of reactions can begin and end at the same time, sharing the identical amplification conditions, and thereby allowing their quantitative analysis and comparison. Using a portable UV lamp and a smartphone camera, we also succeeded in quantifying the amounts of clinically important and human papillomavirus type 16 genes in human serum and SARS-CoV-2's nucleocapsid genes in human serum and saliva, and the limit of detection was as low as 0.1 nM, highly applicable for actual PoC tests in the field with no purification process.

Development of a two-in-one integrated bioassay for simultaneous and rapid on-site detection of Pb2+ and Hg2+ in water

Exposure to heavy metal pollution represents a serious health risk and requires rapid, sensitive, and accurate detection to protect human health. However, the existing methods had various limitations for simultaneous detection of heavy metals. Herein, a novel two-in-one integrated bioassay was constructed for simultaneous on-site determination of Hg2+ and Pb2+ through combining all-fiber bicolor fluorescence biosensor (ABFB), functional nucleic acids (FNAs), and FRET. Hg2+ biosensor, consisting of BQ-T14 and CY-A14, performed its detection based on specifical binding of Hg2+ and T bases to form T-Hg2+-T mismatch structure. GR-5 DNAzyme, consisting of GR-5S and GR-5E, is applied for Pb2+ biosensor. The higher concentration of Hg2+ and Pb2+ results in the more T-Hg2+-T mismatch structure formed and the GR-5E to cleave the more GR-5S, respectively, thus allowing higher detectable fluorescence intensities in two signal channels. Thanks to high selectivity of two FNAs-biosensors and excellent time-resolved effect of the ABFB, the Hg2+ and Pb2+ could be directly and simultaneously quantitated in 12 min, and had the LODs of 4.22 nM and 1.55 nM, respectively. The satisfied recovery rate of real samples was verified by ICP-OES. Our strategy could serve as a simple and cost-effective on-site high-frequency detection of heavy metal ions.

DNAzyme-based sensing probe protected by DNA tetrahedron from nuclease degradation for the detection of lead ions

Lead poisoning endangers soil, plants and human health due to its toxic effect. It is urgent to develop ideal tool for the in vivo detection of Pb2+.In this study, tetrahedron-based Pb2+-sensitive DNAzyme sensor (TPS) is constructed by taking advantages of a classic Pb2+-dependent GR-5 DNAzyme and DNA tetrahedral structure, where the cleavage substrate and DNAzyme are modified with fluorophore FAM and quencher BHQ-1, respectively. DNA tetrahedron is arranged at the terminus of substrate/DNAzyme duplex to offer the protective shield against the nuclease attack. In the absence of Pb2+, FAM and BHQ-1 are kept close and FAM fluorescence is efficiently quenched. However, in the presence of Pb2+ cofactor, the DNAzyme exhibits the catalytic activity and cleaves the substrate strands, spatially separating the FAM away from BHQ-1 and releasing fluorescence. Utilizing the sensing probe, the Pb2+ can be quantitatively detected down to 1 nM without the interference from nontarget metal ions. Even if incubating in the human serum solution for 12 h, no substantial nuclease degradation is detected. In different complex biological milieu, the TPS can preserve the 85% of fluorescence signal, indicating that the developed TPS is a promising tool for the future application in the in vivo detection of Pb2+.

Ultrasensitive electrochemical detection of Pb2+ based on DNAzyme coupling with exonuclease III –Assisted target recycling

A sensitive and selective electrochemical Pb2+ sensor was developed based on GR-5 DNAzyme and exonuclease III-assisted target recycling amplification. When target Pb2+ existed, GR-5 DNAzyme was cleaved into two free DNA fragments. One of the fragments could open the hairpin DNA (HP1) modified on the electrode to form DNA duplex with blunt end at the 3′-terminus and triggered Exo III cleavage process. A large amount of residual part of HP1 were obtained by the Exo III-aided recycling which could hybridized with another hairpin DNA labeled with methylene blue, resulting in the dramatic increase of current. The electrochemical signal of methylene blue positive correlated with the Pb2+ concentration. The proposed sensor has the concentration ranging from 10−11 to 8×10−6 mol/L with a detection limit of 2.74 p mol/L. Moreover, the analysis of Pb2+ concentration in real water revealed that the proposed biosensor has a promising application in environment monitoring.

A label-free lead(II) ion sensor based on surface plasmon resonance and DNAzyme-gold nanoparticle conjugates

Detection of lead(II) (Pb2+) ions in water is important for the protection of human health and environment. The growing demand for onsite detection still faces challenges for sensitive and easy-to-use methods. In this work, a novel surface plasmon resonance (SPR) biosensor based on GR-5 DNAzyme and gold nanoparticles (AuNPs) was developed. Thiolated DNAzyme was immobilized on the gold surface of the sensor chip followed by anchoring the substrate-functionalized AuNPs through the DNAzyme-substrate hybridization. The coupling between the localized surface plasmon (LSP) of AuNPs and the surface plasmon polaritons (SPP) on the gold sensor surface was used to improve the sensitivity. The substrate cleavage in the presence of Pb2+ ions was catalyzed by DNAzyme, leading to the removal of AuNPs and the diminished LSP-SPP coupling. The optimal detection limit was 80pM for the sensor fabricated with 1μM DNAzyme, corresponding to two or three orders of magnitude lower than the toxicity levels of Pb2+ in drinking water defined by WHO and USEPA. By tuning the surface coverage of DNAzyme, the sensitivity and dynamic range could be controlled. This sensor also featured high selectivity to Pb2+ ions and simple detection procedure. Successful detection of Pb2+ ions in groundwater indicates that this method has the prospect in the onsite detection of Pb2+ ions in water. Given the variety of AuNPs and metal-specific DNAzymes, this detection strategy would lead to the development of more sensitive and versatile heavy metal sensors. Graphical abstract.

Cascade-Amplified Microfluidic Particle Accumulation Enabling Quantification of Lead Ions through Visual Inspection

Exposure to lead in drinking water elevates blood lead levels and causes permanent impairment to human body. To provide a preventive measure for lead intoxication, here we report a sensitive visual quantification of lead in drinking water and whole blood. GR-5 DNAzyme can be cleaved by Pb2+, releasing an oligonucleotide T that triggers catalytic assembly of hairpin complex H1H2 connecting magnetic microparticles (MMPs) and polystyrene microparticles (PMPs). After loading the particle solution into a capillary-driven microfluidic device, MMPs-H1H2-PMPs are first attracted and removed by a magnetic separator, and the remaining free PMPs continue flowing along the microchannel until accumulating at a particle dam. As such, more lead ions cause shorter PMP accumulation quantifiable by the naked eye. The method achieved a limit of detection (LOD) of 246 pM, and is extremely selective (> 40,000 folds to other metal ions), and highly tolerant to acidity/basicity (6 ‒ 8.5) and water hardness (55 ‒ 318.3 mg/L). More importantly, high recovery rate (> 78%) in tap water and LOD of 2.57 nM for lead level in whole blood were achieved, demonstrating a visual quantification method for screening water safety and lead intoxication with user-friendly interface.

Combining whispering gallery mode optofluidic microbubble resonator sensor with GR-5 DNAzyme for ultra-sensitive lead ion detection

Lead ions are deleterious pollutants that often reach drinking water, and can cause significant harm to humans (particularly children). An ultra-sensitive lead ion detection method using a whispering gallery mode (WGM) optofluidic microbubble resonator and the classic GR-5 DNAzyme is proposed in this paper. With the auxiliary piranha and Ploy-l-lysine solution, GR-5 DNAzyme was successfully modified on the inner wall of a microbubble. The mode field distribution of the microbubble was analysed, and the optofluidic sensor with thin wall exhibited a maximum bulk refractive index sensitivity of 265.2 nm/RIU. Lead ions at concentrations ranging from 0.1 pM to 100 pM were tested using the proposed WGM optofluidic sensor. The noise was decreased to 2.43 fM using the self-referenced differential method. Thus, a limit of approximately 15 fM was obtained for the detection of lead ions using the WGM optofluidic biosensor. Eight competing metal ions were also used to evaluate the selectivity of the proposed sensor, with results indicating that it has high selectivity for lead ions. Finally, the sensor performance is verified using real samples.

A rapid and colorimetric biosensor based on GR-5 DNAzyme and self-replicating catalyzed hairpin assembly for lead detection

A rapid and colorimetric biosensor for Pb2+detection has been constructed on the basis of Pb2+-dependent GR-5 DNAzyme and the self-replicating catalyzed hairpin assembly (SRCHA) reaction.

Visual detection of lead ions based on nanoparticle-amplified magnetophoresis and Mie scattering

Lead poisoning in drinking water is a serious threat to public health as it may cause permanent damage to the human nervous system and developmental disorders in children. Many detection methods were developed based on fluorometric, electrochemical, and chemiluminescent assays, but they require precise instruments for signal quantification, which limit the practicality in resource-limited sites. Here we provide a visual detection of lead ions based on nanoparticle-amplified magnetophoresis and Mie scattering. Lead ions first react with GR-5 DNAzyme to release a DNA fragment that can connect magnetic microparticles (MMPs) and gold nanoparticles (AuNPs) through DNA hybridization. Next, the connected AuNPs are collected by magnetic separation and then released. Benefit from the hundreds of oligonucleotides on the AuNPs, tiny amount of AuNPs, which represent low concentration of lead ions, can effectively connect a second pair of MMPs and polystyrene microparticles (PMPs). Upon magnetophoresis, it changes solution turbidity from milky to clear because of the reduction of Mie scattering when less free PMPs suspend in the solution. Our results achieved a limit of detection (LOD) at 55 pM in 100 μl (reaction concentration) and 1.108 nM in 5 μl (sample concentration), which is much lower than the prescribed level in drinking water suggested by Environmental Protection Agency (72 nM). In addition, it also achieves high selectivity against other metal ions (> 50,000 to 1) and is compatible with detection in tap water, providing a visual, sensitive detection without complicated instrument.