CRISPR-Cas-based biosensors for the detection of cancer biomarkers.

Along with discovering cancer biomarkers, non-invasive detection methods have played a critical role in early cancer diagnosis and prognostic improvement. Some traditional detection methods have been used for detecting cancer biomarkers, but they are time-consuming and involve materials and human costs. With great flexibility, sensitivity and specificity, the clustered regularly interspaced short palindromic repeats (CRISPR)-associated system provides a wide range of application prospects in this field.

Colorimetric detection of single-nucleotide mutations based on rolling circle amplification and G-quadruplex-based DNAzyme.

In this work, we proposed a sensitive and selective colorimetric assay for single nucleotide mutation (SNM) detection combining rolling circle amplification (RCA) and G-quadruplex/hemin DNAzyme complex formation. In the detection principle, the first step involves ssDNA hybridization with a padlock probe (PLP) DNA, which can discriminate a single base mismatch. The successful ligation is followed by an RCA event to generate an abundance of G-quadruplexes (GQ-RCA) which are then transformed into a DNAzyme (G-quadruplex/hemin complex) by the addition of hemin.

Functional Nucleic Acid Probes Based on Two-Photon for Biosensing.

Functional nucleic acid (FNA) probes have been widely used in environmental monitoring, food analysis, clinical diagnosis, and biological imaging because of their easy synthesis, functional modification, flexible design, and stable properties. However, most FNA probes are designed based on one-photon (OP) in the ultraviolet or visible regions, and the effectiveness of these OP-based FNA probes may be hindered by certain factors, such as their potential for photodamage and limited light tissue penetration. Two-photon (TP) is characterized by the nonlinear absorption of two relatively low-energy photons of near-infrared (NIR) light with the resulting emission of high-energy ultraviolet or visible light.

A turn-on fluorescence strategy for hypochlorous acid detection based on DNAzyme-assisted cyclic signal amplification.

Hypochlorous acid (HClO) is a crucial active oxygen component and one of the innate immunity's barrier substances in the body. Abnormal fluctuations in HClO concentration can lead to increased oxidative stress, cellular dysfunction, and the onset of various diseases. Thus, developing convenient, affordable, efficient, and sensitive methods to monitor HClO concentration in healthcare and pathophysiology research is highly significant.

Aptamer-based expansion microscopy platform enables signal-amplified imaging of dendritic spines.

Super-resolution imaging of dendritic spines (DS) can provide valuable information for mechanistic studies related to synaptic physiology and neural plasticity, but challenged by their small dimension (50-200 nm) below the spatial resolution of conventional optical microscopes. In this work, by combining the molecular recognition specificity of aptamer with high programmability of DNA nanotechnology, we developed an expansion microscopy (ExM) platform for imaging DS with enhanced spatial resolution and amplified signal output. Our results demonstrated that the aptamer probe could specifically bind to DS of primary hippocampal neurons.

A Fluorescent Biosensor for Streptavidin Detection Based on Double-Hairpin DNA-Templated Copper Nanoparticles.

In this paper, we developed a sensitive, label-free and facile fluorescent strategy for detecting streptavidin (SA) based on double-hairpin DNA-templated copper nanoparticles (CuNPs) and terminal protection of small molecule-linked DNA. Herein, a special DNA hairpin probe was designed and synthesized, which contained two poly T single-stranded loops and a nick point in the middle of the stem. Inspired by the concept of the terminal protection interaction, the specific binding of SA to the biotinylated DNA probe can prevent the exonuclease degradation and keep the integrity of DNA probe, which can be used for synthesizing fluorescent CuNPs as a template.

A label-free ThT-assisted fluorescence detection strategy of alkaline phosphatase activity based on MnO nanosheets.

Alkaline phosphatase (ALP) is a metalloenzyme, the level of which is clinically significant as an abnormality of ALP activity results in several diseases. In the present study, we introduced a MnO nanosheet-based assay for ALP detection employing the adsorption and reduction characteristics of G-rich DNA probes and ascorbic acid (AA), respectively. Ascorbic acid 2-phosphate (AAP) was utilized to act as a substrate for ALP which hydrolyzes AAP generating AA.

Recent Advances in Detection for Breast-Cancer-Derived Exosomes.

Breast cancer is the most common malignant tumor in women, its incidence is secret, and more than half of the patients are diagnosed in the middle and advanced stages, so it is necessary to develop simple and efficient detection methods for breast cancer diagnosis to improve the survival rate and quality of life of breast cancer patients. Exosomes are extracellular vesicles secreted by all kinds of living cells, and play an important role in the occurrence and development of breast cancer and the formation of the tumor microenvironment. Exosomes, as biomarkers, are an important part of breast cancer fluid biopsy and have become ideal targets for the early diagnosis, curative effect evaluation, and clinical treatment of breast cancer.

A turn-on fluorescent strategy for alkaline phosphatase detection based on enzyme-assisted signal amplification.

As a representative biochemical indicator, alkaline phosphatase (ALP) is of great importance in indicating and diagnosing clinical diseases. Herein, we developed a signal-on fluorescence sensing method for sensitive ALP activity detection based on the enzyme-assisted target recycling (EATR) technique. In this method, a two-step signal amplification process is designed.

A Label-Free Fluorescence Aptasensor Based on G-Quadruplex/Thioflavin T Complex for the Detection of Trypsin.

A novel, label-free fluorescent assay has been developed for the detection of trypsin by using thioflavin T as a fluorescent probe. A specific DNA aptamer can be combined by adding cytochrome c. Trypsin hydrolyzes the cytochrome c into small peptide fragments, exposing the G-quadruplex part of DNA aptamer, which has a high affinity for thioflavin T, which then enhances the fluorescence intensity.

Detection of DNA Methyltransferase Activity via Fluorescence Resonance Energy Transfer and Exonuclease-Mediated Target Recycling.

In this study, a sensitive method for detecting DNA methyltransferase (MTase) activity was developed by combining the effective fluorescence resonance energy transfer (FRET) of cationic conjugated polymers and exonuclease (Exo) III-mediated signal amplification. DNA adenine MTase targets the GATC sequence within a substrate and converts the adenine in this sequence into N6-methyladenine. In the method developed in this study, the methylated substrate is cleaved using Dpn I, whereby a single-stranded oligodeoxynucleotide (oligo) is released.

A fluorescence strategy for the rapid detection of miRNA-21 based on G-quadruplex and cyclic amplification signal.

MicroRNAs (miRNAs) are commonly used as biomarkers for the diagnosis of tumors. Since miRNA expression is strongly correlated to carcinogenesis, the detection of miRNA concentration in cells would be valuable for the diagnosis and evaluation of tumors. In this study, we proposed a system using two strands of DNA, one modified by a phosphate group at the 5' end, called Cap, and the other with a hairpin structure, called HP.

Label-Free miRNA-21 Analysis Based on Strand Displacement and Terminal Deoxynucleotidyl Transferase-Assisted Amplification Strategy.

MicroRNAs (miRNAs) are regarded as a rising star in the biomedical industry. By monitoring slight increases in miRNA-21 levels, the possibilities of multi-type malignancy can be evaluated more precisely and earlier. However, the inconvenience and insensitivity of traditional methods for detecting miRNA-21 levels remains challenging.

A fluorescent assay for alkaline phosphatase activity based on phosphorylation protection and DNAzyme-assisted amplification.

Alkaline phosphatase is one of the most important tool enzymes and diseases indicator, monitoring ALP activity with convenient, precise, efficient and sensitive methods plays a fundamental role in modern life and healthcare industries. In this study, we described a novel method for ALP analysis based on Pb dependent DNAzyme. By modifying DNAzyme sequence with terminal phosphate group and introducing exonuclease I (exo I), we managed to analyze ALP by utilizing its causal function of DNAzyme probe from exo I mediated degradation and function of triggering the subsequent cleavage of the hairpin reporting probe.

Recent advances in biosensor for DNA glycosylase activity detection.

Base excision repair (BER) is vital for maintaining the integrity of the genome under oxidative damage. DNA glycosylase initiates the BER pathway recognizes and excises the mismatched substrate base leading to the apurinic/apyrimidinic site generation, and simultaneously breaks the single-strand DNA. As the aberrant activity of DNA glycosylase is associated with numerous diseases, including cancer, immunodeficiency, and atherosclerosis, the detection of DNA glycosylase is significant from bench to bedside.

A Label-Free Fluorometric Glutathione Assay Based on a Conformational Switch of G-quadruplex.

In this paper, a label-free fluorescent method for glutathione (GSH) detection based on a thioflavin T/G-quadruplex conformational switch is developed. The sensing assay is fabricated depending on the virtue of mercury ions to form a thymine-thymine mismatch, which collapses the distance between two ssDNA and directs the guanine-rich part to form an intra-strand asymmetric split G-quadruplex. The newly formed G-quadruplex efficiently reacts with thioflavin T and enhances the fluorescent intensity.

An Exonuclease I-Aided Turn-Off Fluorescent Strategy for Alkaline Phosphatase Assay Based on Terminal Protection and Copper Nanoparticles.

As an important DNA 3'-phosphatase, alkaline phosphatase can repair damaged DNA caused by replication and recombination. It is essential to measure the level of alkaline phosphatase to indicate some potential diseases, such as cancer, related to alkaline phosphatase. Here, we designed a simple and fast method to detect alkaline phosphatase quantitively.

A signal-on fluorescence-based strategy for detection of microRNA-21 based on graphene oxide and λ exonuclease-based signal amplification.

MicroRNA (miRNA) expression is perturbed in various diseases. Herein, we have aimed to develop a novel and rapid fluorescence-based assay for detecting microRNA-21 (miR-21) activity based on FAM molecular signal amplification and graphene oxide (GO) quenching. In this system, a single stranded DNA (ssDNA) with a phosphate group at the 5'-end is labeled with a FAM molecular label at the 3'-end.

Detection of Streptavidin Based on Terminal Protection and Cationic Conjugated Polymer-Mediated Fluorescence Resonance Energy Transfer.

In this paper, a fast and simple strategy for sensitive detection of streptavidin (SA) was proposed based on terminal protection of small molecule-linked DNA and cationic conjugated polymer-mediated fluorescence resonance energy transfer (FRET). In principle, we designed a biotin-labelled DNA probe (P1) as the recognitive probe of SA, along with a complementary DNA probe (P2) to form double-stranded DNA (dsDNA) with P1. SYBR Green I (SG I) as a fluorescent dye was further used to specifically bind to dsDNA to emit stronger fluorescence.

Sensitive Fluorescence Assay for the Detection of Alkaline Phosphatase Based on a Cu-Thiamine System.

The authors describe a novel, facile, and sensitive fluorometric strategy based on a Cu-thiamine (Cu-TH) system for the detection of alkaline phosphatase (ALP) activity and inhibition. The principle of the method is as follows. Under a basic conditions, TH, which does not exhibit a fluorescence signal, is oxidized into fluorescent thiochrome (TC) by Cu.

Amplified Fluorescent Aptasensor for Ochratoxin A Assay Based on Graphene Oxide and RecJ Exonuclease.

In this study, we developed an aptamer-based fluorescent sensing platform for the detection of ochratoxin A (OTA) based on RecJ exonuclease-assisted signal amplification and interaction between graphene oxide (GO) and the OTA aptamer (OTA-apt). After optimizing the experimental conditions, the present aptamer-based sensing system can exhibit excellent fluorescent response in the OTA assay, with a limit of detection of 0.07 ng/mL.

A novel fluorescent assay based on DNAzyme-assisted detection of prostate specific antigen for signal amplification.

Prostate specific antigen (PSA) is one of the most common biomarkers for the management of prostate cancer. However, it still remains urgent to develop highly sensitive, cost-effective and selective strategies for PSA assay. In this paper, we developed a low-cost, highly sensitive and specific analytical strategy for the detection of PSA by using a fluorescence sensor based on Pb-dependent DNAzyme.

A turn-on fluorescence assay of alkaline phosphatase activity based on an enzyme-triggered conformational switch of G-quadruplex.

Herein, a turn-on fluorescence assay was introduced for alkaline phosphatase (ALP) detection based on ThT/G-quadruplex system. The basis of the method is that chelation of guanine bases at the binding sites by Ag blocks G-quadruplex formation and decreases the fluorescence intensity sharply. In the presence of ALP, ascorbic acid 2-phosphate (AAP) is hydrolyzed to form ascorbic acid (AA) which in turn reduces Ag to Ag.

Label-free one-step fluorescent method for the detection of endonuclease activity based on thioflavin T/G-quadruplex.

Endonucleases, one of the basic tool enzymes of modern molecular biology and medical genetics, have also been clarified as the potential targets for antimicrobial and antiviral drugs screening. However, traditional assays to monitor endonuclease activity can be expensive, time-consuming, or laborious. In order to provide new detective platform, we proposed a novel label-free one-step fluorescent method for the detection of endonuclease activity based on cleavage-induced G-quadruplex formation.