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The traditional detection of telomerase activity is mainly based on the polymerase chain reaction (PCR), which has the disadvantages of being time-consuming and susceptible to interferences; thus, here, we propose a facile method for the fabrication of fluorescent tungsten oxide quantum dots (WO x QDs) and employ them for telomerase activity sensing. It is found that the fluorescence of WO x QDs can be significantly quenched by hemin based on the inner filter effect (IFE). However, in the presence of telomerase, the primer-DNA can be extended to generate repeating units of TTAGGG to form G-quadruplex and thus, hemin can be encapsulated to reduce its absorbance, resulting in decreased IFE and efficient fluorescence recovery of WO x QDs. Based on the fluorescence changes of IFE between hemin and WO x QDs, the telomerase activity within the range of 50-30 000 HeLa cells can be detected and the lowest detection amount can reach 17 cells. The method exhibits good versatility that can also be applied to telomerase detection in A549 and L929 cells. In addition, because of the good biocompatibility of the sensor, it can be used for the real-time monitoring of telomerase activity in living cells, thus showing great potential in tumor diagnosis and inhibitor drug screening.

A ratiometric fluorometric method is described for the determination of arsenate via its inhibitory effect on the activity of the enzyme acid phosphatase. A nanoprobe was designed that consists of CdSe/ZnS quantum dots (QDs) coated with the terbium(III) complex of guanosine monophosphate (Tb-GMP). The nanoprobe was synthesized from carboxylated QDs, Tb(III) and GMP via binding of Tb(III) by both the carboxy and the phosphate groups. The nanoprobe, under single-wavelength excitation (at 280 nm), displays dual (red and green) emission, with peaks at around 652 nm from the QDs, and at 547 nm from the Tb-GMP coordination polymers. It is shown to be a viable nanoprobe for fluorometric determination of As(V) detection through it inhibitory action on the activity of acid phosphatase (ACP). The enzyme destroys the Tb-GMP structure via hydrolysis of GMP, and hence the fluorescence of the Tb-GMP complex is quenched. In contrast, the fluorescence of the CdSe/ZnS QDs remains inert to ACP. It therefore can serve as an internal reference signal. In the presence of arsenate (an analog of phosphate), the activity of ACP is inhibited due to competitive binding. Thus, hydrolysis of GMP is prevented. These findings were used to design a ratiometric fluorometric method for the quantification of As(V). The ratio of fluorescences at 547 and 652 nm increases linearly in the 0.5 to 200 ppb As(V) concentation range, and the limit of detection is 0.39 ppb. Under a UV lamp, the probe shows distinguishable color from green to red on increasing the concentration of As(V). Graphical abstract Schematic illustration of CdSe/ZnS quantum dot coated with carboxy-PEG and modified with the terbium(III)-GMP complex as a fluorescent nanoprobe for ratiometric determination of arsenate via its inhibition of ACP activity.

Mercuric ion (Hg II ) is a stable form of mercury pollution with high toxicity and bioaccumulation ability, and its sensitive and visible determination is of great importance. Herein, a simple method to significantly improve the complexation reaction rate between porphyrin and manganous ions (Mn II ) has been implemented by using the synergistic effect of trace nitrogen-doped graphene quantum dots (NGQDs) and Hg(II). A mechanism is proposed in accordance with a substitution reaction in which the deformed porphyrin nucleus by relatively larger Hg(II) ions is favorable for attacking small divalent metal ions carried by NGQDs from the back. Meanwhile, the formation of metalloporphyrin is accompanied by the absorption red-shift and fluorescence quenching of porphyrins; simultaneously, the fluorescence of NGQDs is gradually enhanced because of the inner filter effect between porphyrins and NGQDs. Thus, the ratiometric fluorescence and colorimetric methods for trace Hg II sensing have been proposed on the basis of the distinct absorption/fluorescence spectral changes, which have potential application in complex environmental and biological conditions.

Herein, a novel electrochemiluminescence resonance energy transfer (ECL-RET) biosensor using graphene quantum dots (GQDs) as donor and graphene oxide (GO) as acceptor for monitoring the activity of protein kinase was presented for the first time. Anti-phosphoserine antibody conjugated graphene oxide (Ab-GO) nonocomposite could be captured onto the phosphorylated peptide/GQDs modified electrode surface through antibody-antigen interaction in the presence of casein kinase II (CK2) and adenosine 5'-triphosphate (ATP), resulting in ECL from the GQDs quenching by closely contacting GO. This ECL quenching degree was positively correlated with CK2 activity. Therefore, on the basis of ECL-RET between GQDs and GO, the activity of protein kinase can be detected sensitively. This biosensor can also be used for quantitative analysis CK2 activity in serum samples and qualitative screening kinase inhibition, indicating the potential application of the developed method in biochemical fundamental research and clinical diagnosis., (Copyright © 2015 Elsevier B.V. All rights reserved.)

A novel Au NP mediated dual-potential ECL ratiometric approach for highly sensitive protein kinase activity and inhibition assay has been developed based on the simultaneous decrease of cathodic ECL from GQDs and enhancement of anodic ECL from luminol in the same bioanalysis.

In this work, we designed a novel detection strategy to realize simultaneous determination of multiplex lectin by labeling glucosamine (G1) and galactosamine (G2) with different-colored semiconductor quantum dots (QDs). On the basis of the agglutination of the aminosugar-labeled QDs induced by the exclusive binding between the lectin and sugar on the QDs surfaces, the fluorescence emission of the QDs supernatant after centrifugation decreased with relevant lectin concentration [i.e., when concanavalin A (Con A) exists alone], only green color fluorescence emission from QDs-G1 supernatant decreased, so it is peanut agglutinin (PNA) and red color fluorescence emission from QDs-G2. Moreover, since QDs can be simultaneously excited with multiple fluorescence colors and have a larger Stokes shift than organic fluorophores, when both Con A and PNA are present in the sample, both of the green and red color fluorescence emission from QDs-G1 and QDs-G2 supernatant would decrease, thus realizing the simultaneous determination of Con A and PNA. The detection limits of Con A and PNA are 0.30 and 0.18 nM (3σ), respectively. Furthermore, the present detection method not only can determine the protein/lectins by fluorescence spectral method but also can realize visualization detection by UV lamp illumination. To the best of our knowledge, this is the first report of such analytical method in multiple and simultaneous lectin detection.

A simple and sensitive photoluminescence (PL) assay for the activity of a protein kinase based on the selective aggregation of phosphorylated peptide-graphene quantum dot (GQD) conjugates triggered by Zr(4+) ion coordination has been established. With more sophisticated design of the peptide substrate sequences, detecting other enzymes could also be possible. Under optimal conditions, a linear relationship between the decreased PL intensity of peptide-GQD conjugates and the concentration of casein kinase II (CK2) in the range from 0.1 to 1.0 unit mL(-1) with a detection limit of 0.03 unit mL(-1) (3σ) was obtained. The EC50 value (i.e., the enzyme concentration producing 50% substrate conversion) for CK2 was evaluated to be 0.34 unit mL(-1). The proposed method showed potential applications in kinase inhibitor screening. To demonstrate the potential of this GQD-based platform for screening of kinase inhibitors in real biological systems, the inhibition of CK2 phosphorylation activity by four different inhibitors (ellagic acid, 5,6-dichlorobenzimidazole-l-β-d-ribofuranoside, emodin, and quercetin) was tested in human serum by comparing signals from samples incubated with the inhibitors against that without any inhibitor. As expected, in the presence of inhibitors, the PL intensity increased with increasing inhibitor efficiency. The IC50 value (inhibitor concentration producing 50% inhibition) for ellagic acid was estimated to be 0.041 μM. The developed protocol provides a new and promising tool for the analysis of both the enzyme and its inhibitors with low cost and excellent performance.

Using graphene quantum dots (GQDs) with a boronic acid-substituted bipyridinium salt (BBV), a label-free fluorescence assay for glucose detection is presented.