Jia Ge

A label-free method for detecting biothiols based on poly(thymine)-templated copper nanoparticles.

A novel label-free nanosensor has been developed for detecting biothiols including cysteine, glutathione and homocysteine based on poly(thymine)-templated fluorescent copper nanoparticles (CuNPs), which were controlled through thymine-Hg(II)-thymine coordination. This assay provides a simple, cost-effective, and sensitive platform for the detection of biothiols. By employing this turn-on sensor, biological thiols, such as cysteine, glutathione and homocysteine, are successfully detected at concentrations as low as 12.5nM for cysteine, 15nM for glutathione, and 20nM for homocysteine. The sensing system also exhibits high selectivity against other amino acids, and the application of the sensor for biological fluids shows that the proposed method works well for real samples.

An aptamer-based signal-on bio-assay for sensitive and selective detection of Kanamycin A by using gold nanoparticles.

In this study, a simple and sensitive aptamer-based fluorescence method for the detection of Kanamycin A by using gold nanoparticles (AuNPs) has been developed. In this assay, AuNPs were utilized as DNA nanocarrier as well as efficient fluorescence quencher. In the absence of Kanamycin A, dye-labeled aptamer could be adsorbed onto the surface of AuNPs and the fluorescence signal was quenched. In the presence of Kanamycin A, the specific binding between dye-labeled aptamer and its target induced the formation of rigid structure, which led to dye-labeled aptamer releasing from the surface of AuNPs and the fluorescence intensity was recovered consequently. Under optimum conditions, calibration modeling showed that the analytical linear range covered from 0.8nM to 350nM and the detection limit of 0.3nM was realized successfully. This proposed bio-assay also showed high selectivity over other antibiotics. Meanwhile, this strategy was further used to determine the concentrations of Kanamycin A in milk sample with satisfying results.

Nitrogen-doped Carbon Dots Mediated Fluorescent on-off Assay for Rapid and Highly Sensitive Pyrophosphate and Alkaline Phosphatase Detection

In this report, a novel fluorescent sensing platform using nitrogen-doped carbon dots (N-CDs) as probes for fluorescence signal transmission has been designed for the detection of significant biomolecules pyrophosphate (PPi) and alkaline phosphatase (ALP). The high fluorescent N-CDs could be selectively quenched by Cu2+, and recovered by the addition of PPi because PPi preferentially binds to Cu2+. Once ALP was introduced into the system, ALP can specifically hydrolyze PPi into Pi, the intense fluorescence of N-CDs could be quenched again due to the recombination of the as-released Cu2+ with N-CDs. So, fluorescence of N-CDs is regulated by an ALP-triggered reaction. Based on this strategy, we demonstrated that N-CDs could serve as a very effective fluorescent sensing platform for label-free, sensitive and selective detection of PPi and ALP with low detection limit of 0.16 μM and 0.4 U/L for PPi and ALP, respectively. Moreover, the assay time is just around 0.5 min for PPi and 30 min for ALP. This developed strategy shows remarkable advantages including sensitive, rapid, simple, convenient, and low-cost and so forth. Furthermore, this method was also successfully applied to monitor ALP in human serum, which indicates its great potential for practical applications in biological and clinical diagnosis.

PSS-GN nanocomposites as highly-efficient peroxidase mimics and their applications in colorimetric detection of glucose in serum

In this report, poly(sodium styrene sulfonate)-functionalized graphene nanosheets (PSS-GN) with good water dispersibility were prepared and for the first time proposed as a peroxidase-like mimic. The intrinsic catalytic activity of PSS-GN nanocomposites was investigated and the results showed that PSS-GN nanocomposites had higher mimetic enzyme catalytic activity compared with horseradish peroxidase (HRP), which might be due to the stronger binding affinity between negatively charged PSS and the positively charged peroxide substrate (3,3,5,5-tetramethylbenzidine, TMB). On the basis of the high catalyze activity, these PSS-GN nanocomposites were utilized for sensitive and selective colorimetric detection of H2O2 as well as glucose. The linearity between analyte concentration and system absorption ranged from 0.005 to 1 mM for H2O2 and 0.006 to 0.4 mM for glucose with a detection limit of 0.15 μM and 0.28 μM, respectively. Moreover, this strategy was further utilized to determine the concentrations of glucose in serum samples with satisfying results. This assay exhibits good analytical performance, acceptable stability and good selectivity. We envision that this sensing system could be used as a tool for a wide range of potential applications in enzyme-based chemical and biological analysis.

A novel label-free fluorescent molecular beacon for the detection of 3′–5′ exonuclease enzymatic activity using DNA-templated copper nanoclusters

The 3′–5′ exonuclease plays key roles in maintaining genome stability; therefore, the detection of 3′–5′ exonuclease activity is very important for disease diagnosis and drug development. In this study, we present a novel highly sensitive and label-free fluorescent assay to detect the activity of exonuclease III (Exo III) using molecular beacon DNA with a polythymine (T) loop as a template for the formation of fluorescent copper nanoparticles (CuNPs). Upon the addition of Exo III, the molecular beacon probe was digested into mono- or short oligonucleotides by the exonucleases and failed to form fluorescent CuNPs. As a result, the fluorescence intensity of CuNPs decreased. This method exhibited a very low detection limit of 0.02 U mL−1 for Exo III. The designed molecular beacon probe can be a potential label-free strategy for the detection of other nuclease activity.

Sensitive and label-free T4 polynucleotide kinase/phosphatase detection based on poly(thymine)-templated copper nanoparticles coupled with nicking enzyme-assisted signal amplification

Dephosphorylation of DNA by DNA 3′-phosphatases plays significant regulatory roles in DNA damage repair, replication, and recombination. Here, we present a novel, highly sensitive and label-free fluorescent assay for T4 polynucleotide kinase/phosphatase (T4 PNKP) activity and its inhibitors on the basis of nicking enzyme-assisted signal amplification and poly(thymine)-templated fluorescent copper nanoparticles (CuNPs) as a fluorescent indicator. It was designed such that an A-rich hairpin primer with a 3′-phosphoryl end can serve as the substrate for T4 PNKP. Once the phosphorylated hairpin primer was hydrolyzed by T4 PNKP, the resulting hairpin primer with a 3′-hydroxyl end was immediately elongated to form a long double-strand product using DNA polymerase as well as a double-stranded recognition site for nicking enzyme Nb.BbvCI. With the help of nicking enzyme Nb.BbvCI, the double-strand product could be cleaved to release a single-stranded poly-(thymine) (poly T) DNA, which could be used as a template for the formation of fluorescent CuNPs. Subsequently, the remaining probe could be reacted as the primer to trigger the next cycle of polymerization and scission process, thus significantly improving the sensitivity of the sensor. Furthermore, the application of the proposed strategy for screening T4 PNKP inhibitors also demonstrated satisfactory results. With the advantages mentioned above, this simple, cost-effective, and highly sensitive strategy has potential for the study of DNA damage repair mechanisms.

Synthesis of Luminescent Carbon Dots with Ultrahigh Quantum Yield and Inherent Folate Receptor-Positive Cancer Cell Targetability

Carbon dots (CDs) have a wide range of applications in chemical, physical and biomedical research fields. We are particularly interested in the use of CDs as fluorescence nanomaterials for targeted tumor cell imaging. One of the important aspects of success is to enhance the fluorescence quantum yields (QY) of CDs as well as increase their targetability to tumor cells. However, most of the reported CDs are limited by relative low QY. In the current study, for the first time, one-step synthesis of highly luminescent CDs by using folic acid (FA) as single precursor was obtained in natural water through hydrothermal method. The as-prepared CDs exhibited QY as high as 94.5% in water, which is even higher than most of organic fluorescent dyes. The obtained CDs showed excellent photoluminescent activity, high photostability and favorable biocompatibility. The FA residuals in CDs led to extraordinary targetability to cancer cells and promoted folate receptor-mediated cellular uptake successfully, which holds a great potential in biological and bioimaging studies.

A label-free aptasensor for highly sensitive ATP detection by using exonuclease I and oligonucleotide-templated fluorescent copper nanoparticles

In this work, we developed a simple and highly sensitive fluorescence method for the determination of ATP based on a designed multifunctional oligonucleotide, which holds the ability of synthesizing fluorescent copper nanoparticles (CuNPs) as well as recognizing ATP molecules. In the absence of ATP, this multifunctional probe containing an ATP aptamer and poly T strand can be digested upon the addition of Exo I, which leads to the failure of synthesizing poly T-templated fluorescent CuNPs, whereas in the presence of ATP, the ATP aptamer folds into a G-quadruplex structure, which makes it resistant to digestion by Exo I. Accordingly, the poly T strand is reserved and can be used as an efficient template for the formation of CuNPs with strong fluorescence. Thus, the concentration of ATP could be identified very easily by observing fluorescence changes of this sensing system. This signal-on assay exhibits high sensitivity to ATP with a detection limit of 500 nM in the range of 1–80 μM. Further specificity investigation and real sample anti-interference experiments show that this assay also has high selectivity and anti-interference ability.

A rapid and sensitive turn-on fluorescent probe for ascorbic acid detection based on carbon dots–MnO2 nanocomposites

Ascorbic acid (AA) is a significant small biomolecule and plays a key role in many biochemical processes. In this work, a rapid turn-on fluorescent probe was developed for sensitive and selective sensing of AA based on carbon dots–MnO2 nanocomposites. The nanocomposites were constructed by in situ synthesis of MnO2 nanosheets in carbon dot (C-dot) solution, and the fluorescence of C-dots was quenched with the formation of the nanocomposites due to the fluorescence resonance energy transfer (FRET) from C-dots to the generated MnO2 nanosheets. Once AA was introduced into the system, MnO2 nanosheets were reduced to Mn2+ and the fluorescence of C-dots would be recovered. Under the optimized conditions, this method shows rapid and sensitive response toward AA, the assay time is 2 min and the limit of detection is 68 nM. Moreover, the carbon dots–MnO2 nanocomposite based fluorescence sensing system was successfully employed to determine AA in commercial tablets with satisfactory results. Taking full advantage of C-dots and MnO2 nanosheets, this probe shows remarkable properties including easy operation, low cost and good biocompatibility, and has great potential to be used in biological and clinical diagnostic applications.

Label-free biosensor based on dsDNA-templated copper nanoparticles for highly sensitive and selective detection of NAD+

We have developed a novel label-free fluorescence approach for nicotinamide adenine dinucleotide (NAD+) detection by coupling poly(AT-TA) dsDNA-templated fluorescent copper nanoparticles (CuNPs) as a fluorescence indicator with a dumbbell DNA probe designed for ligation. Herein, we designed a dumbbell probe which can form a self-complementary structure at both ends. In the absence of NAD+, the extension reaction can be performed from the exposed 3′-end, and instantly opened the hybridized structure at the 5′-end, producing a chimeric poly(AT-TA) double-stranded fragment which can be used as a highly-efficient template for the formation of CuNPs, showing high fluorescence. Whereas in the presence of NAD+, together with the E. coli DNA ligase, the 5′-phosphoryl end and the 3′-end of the DNA probe can be ligated to block the extension reaction, showing slight fluorescence. This novel NAD+ assay is label-free, fast and easy to operate, highly sensitive and selective, which holds wide applicability as a robust analytical tool in NAD+ related biological processes and clinical diagnosis.