Isothermal amplification of nucleic acids is a simple process that rapidly and efficiently accumulates nucleic acid sequences at constant temperature. Since the early 1990s, various isothermal amplification techniques have been developed as alternatives to polymerase chain reaction (PCR). These isothermal amplification methods have been used for biosensing targets such as DNA, RNA, cells, proteins, small molecules, and ions. The applications of these techniques for in situ or intracellular bioimaging and sequencing have been amply demonstrated. Amplicons produced by isothermal amplification methods have also been utilized to construct versatile nucleic acid nanomaterials for promising applications in biomedicine, bioimaging, and biosensing. The integration of isothermal amplification into microsystems or portable devices improves nucleic acid-based on-site assays and confers high sensitivity. Single-cell and single-molecule analyses have also been implemented based on integrated microfluidic systems. In this review, we provide a comprehensive overview of the isothermal amplification of nucleic acids encompassing work published in the past two decades. First, different isothermal amplification techniques are classified into three types based on reaction kinetics. Then, we summarize the applications of isothermal amplification in bioanalysis, diagnostics, nanotechnology, materials science, and device integration. Finally, several challenges and perspectives in the field are discussed.

Isothermal Amplification of Nucleic Acids

Ultra-small fluorescent metal nanoclusters: Synthesis and biological applications

The emergence of graphene nanosheet (GN, 2010 Nobel Prize for Physics) has recently opened up an exciting new field in the science and technology of two-dimensional (2D) nanomaterials with continuously growing academic and technological impetus. GN exhibits unique electronic, optical, magnetic, thermal and mechanical properties arising from its strictly 2D structure and thus has many important technical applications. Actually, GN-based materials have enormous potential to rival or even surpass the performance of carbon nanotube-based counterparts, given that cheap, large-scale production and processing methods for high-quality GN become available. Therefore, the studies on GN in the aspects of chemistry, physical, materials, biology and interdisciplinary science have been in full flow in the past five years. In this critical review, from the viewpoint of chemistry and materials, we will cover recent significant advances in synthesis, molecular engineering, thin film, hybrids, and energy and analytical applications of the “star-material” GN together with discussion on its major challenges and opportunities for future GN research (315 references).

Graphene nanosheet: synthesis, molecular engineering, thin film, hybrids, and energy and analytical applications

Carbon Nanomaterials for Biological Imaging and Nanomedicinal Therapy

The great success of electrochemiluminescence (ECL) for in vitro diagnosis (IVD) and its promising potential in light-emitting devices greatly promote recent ECL studies. More than 45% of ECL articles were published after 2010, and the first international meeting on ECL was held in Italy in 2014. This critical review discusses recent vibrant developments in ECL, and highlights novel ECL phenomena, such as wireless ECL devices, bipolar electrode-based ECL, light-emitting electrochemical swimmers, upconversion ECL, ECL resonance energy transfer, thermoresponsive ECL, ECL using shape-controlled nanocrystals, and ECL as an ion-selective electrode photonic reporter, a paper-based microchip, and a self-powered microfluidic ECL platform. We also comment on the latest progress in bioassays, light-emitting devices and, the computational approach for the ECL mechanism study. Finally, perspectives and key challenges in the near future are addressed (198 references).

Recent advances in electrochemiluminescence

Amplified Aptamer-Based Assay through Catalytic Recycling of the Analyte†

Electrogenerated chemiluminescence from Au nanoclusters.

In this study, a novel method for the fast, sensitive and selective detection of Cu2+ using gold nanoparticles (AuNPs) was developed and used in immunoassays. In the presence of L-cysteine, L-cysteine can bind to the surface of citrate-stabilized AuNPs through Au-S bonds. As a result, aggregation of AuNPs occurs through electrostatic interactions between the cysteine-bound AuNPs. In contrast, in the presence of Cu2+, Cu2+ can catalyze O2 oxidation of cysteine, leading to the quick formation of disulfide cystine. An increase in the concentration of Cu2+ decreased L-cysteine-induced AuNPs aggregation by decreasing the number of free cysteine thiol groups, and the solution color changed from purple to red. Therefore, the concentration of Cu2+ can be detected with the naked eye or with ultraviolet–visible spectroscopy, and the detection limits of Cu2+ were 20 nM and 10 nM, respectively. This sensitivity was approximately three orders of magnitude higher than that of traditional AuNPs-based colorimetric Cu2+ detection methods. Because of the high sensitivity of the proposed method, we further used it with a labeled antibody in colorimetric immunoassays. The detection limit of the cancer biomarker α-fetoprotein was 2 ng ml−1, which is comparable to the detection limit of the enzyme-linked immunosorbent assay method. Huang-Hao Yang and co-workers at Fuzhou University have devised an ultra-sensitive method to detect copper ions (Cu2+), whose accumulation in the body has been linked to several diseases. The researchers' method relies on gold nanoparticles, popular in colourimetric sensing because they turn a solution from red to purple on aggregation. Here, however, instead of triggering the aggregation directly, the copper ions catalyse a ‘supporting reaction’ – the oxidation of a monomeric amino acid (cysteine) into a dimeric one (cystine). Only cysteine triggers the aggregation of the nanoparticles, which means the presence or absence of copper results in a red or purple solution, respectively. The method's sensitivity is very high because only catalytic quantities of copper are needed. The researchers further used this rapid, convenient, sensitive method in an immunoassay for a human protein. A novel method based on the AuNPs for fast, sensitive and selective detection of Cu2+ was developed and applied in immunoassays.

/pdf/ultrasensitive-detection-of-cu2-with-the-naked-eye-and-1r80v80b0r.pdf

Ultrasensitive detection of Cu2+ with the naked eye and application in immunoassays

Label-free and fluorescence turn-on aptasensor for protein detection via target-induced silver nanoclusters formation.

Nowadays, bacterial infection is very common in the world. As a result of overusing antibiotics, several human pathogenic bacteria have become resistant to most of the clinically approved antibiotics. This has created an urgent need for the development of new methods to kill pathogenic bacteria. Here, we describe a novel method for the selective killing of pathogenic bacteria using antibody functionalized nanoscale reduced graphene oxide (NRGO). The antibody–NRGO complex exhibited little toxicity without near-infrared (NIR) irradiation. However, when irradiated with a NIR laser at a low power density (400 mW cm−2), the antibody–NRGO complex exhibited excellent photothermal properties and killed captured pathogenic bacteria specifically.

Targeted photothermal ablation of pathogenic bacterium, Staphylococcus aureus, with nanoscale reduced graphene oxide

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