Yiping Chen

One-Step Detection of Pathogens and Viruses: Combining Magnetic Relaxation Switching and Magnetic Separation

We report a sensing methodology that combines magnetic separation (MS) and magnetic relaxation switching (MS-MRS) for one-step detection of bacteria and viruses with high sensitivity and reproducibility. We first employ a magnetic field of 0.01 T to separate the magnetic beads of large size (250 nm in diameter) from those of small size (30 nm in diameter) and use the transverse relaxation time (T2) of the water molecules around the 30 nm magnetic beads (MB30) as the signal readout of the immunoassay. An MS-MRS sensor integrates target enrichment, extraction, and detection into one step, and the entire immunoassay can be completed within 30 min. Compared with a traditional MRS sensor, an MS-MRS sensor shows enhanced sensitivity, better reproducibility, and convenient operation, thus providing a promising platform for point-of-care testing.

Surface Modification of Gold Nanoparticles with Small Molecules for Biochemical Analysis

As one of the major tools for and by chemical science, biochemical analysis is becoming increasingly important in fields like clinical diagnosis, food safety, environmental monitoring, and the development of chemistry and biochemistry. The advancement of nanotechnology boosts the development of analytical chemistry, particularly the nanoparticle (NP)-based approaches for biochemical assays. Functional NPs can greatly improve the performance of biochemical analysis because they can accelerate signal transduction, enhance the signal intensity, and enable convenient signal readout due to their unique physical and chemical properties. Surface chemistry is a widely used tool to functionalize NPs, and the strategy for surface modification is of great significance to the application of NP-mediated biochemical assays. Surface chemistry not only affects the quality of NPs (stability, monodispersity, and biocompatibility) but also provides functional groups (-COO-, -NH3+, -CHO, and so on) or charges that can be exploited for bioconjugation or ligand exchange. Surface chemistry also dictates the sensitivity and specificity of the NP-mediated biochemical assays, since it is vital to the orientation, accessibility, and bioactivity of the functionalized ligands on the NPs. In this Account, we will focus on surface chemistry for functionalization of gold nanoparticles (AuNPs) with small organic molecules for biochemical analysis. Compared to other NPs, AuNPs have many merits including controllable synthesis, easy surface modification and high molar absorption coefficient, making them ideal probes for biochemical assays. Small-molecule functionalized AuNPs are widely employed to develop sensors for biochemical analysis, considering that small molecules, such as amino acids and sulfhydryl compounds, are more easily and controllably bioconjugated to the surface of AuNPs than biomacromolecules due to their less complex structure and steric hindrance. The orientation and accessibility of small molecules on AuNPs in most cases can be precisely controlled without compromising their bioactivity as well, thus ensuring the performance, such as the specificity and sensitivity, of AuNP-based biochemical assays. This Account reviews recent progress in the surface chemistry of functionalized AuNPs for biochemical assays. The surface chemistries mainly include click chemistry, ligand exchange reaction, and coordination-based recognition. These surface-modified AuNPs allow for assaying a range of important biochemical markers including metal ions, small biomolecules, enzymes, and antigens and antibodies. Applications of these systems range from environmental monitoring to medical diagnostics. This Account highlights the advantages and limitations (sensitivity, detection efficiency, and stability) that AuNP-mediated assays still have compared with conventional analytical methods. This Account also discusses the future research directions of surface-modified AuNP-mediated biochemical analysis. The main aim of this Account is to summarize the current surface modification strategies for AuNPs and further demonstrate how to make use of surface modification strategies to effectively improve the performance of AuNP-mediated analytical methods for a wide variety of applications relating to biochemical analysis.

Horseradish Peroxidase-Mediated, Iodide-Catalyzed Cascade Reaction for Plasmonic Immunoassays

This report outlines an enzymatic cascade reaction for signal transduction and amplification for plasmonic immunoassays by using horseradish peroxidase (HRP)-mediated aggregation of gold nanoparticles (AuNPs). HRP-catalyzed oxidation of iodide and iodide-catalyzed oxidation of cysteine is employed to modulate the plasmonic signals of AuNPs. It agrees well with the current immunoassay platforms and allows naked-eye readout with enhanced sensitivity, which holds great promise for applications in resource-constrained settings.

Click Chemistry-Mediated Nanosensors for Biochemical Assays

Click chemistry combined with functional nanoparticles have drawn increasing attention in biochemical assays because they are promising in developing biosensors with effective signal transformation/amplification and straightforward signal readout for clinical diagnostic assays. In this review, we focus on the latest advances of biochemical assays based on Cu (I)-catalyzed 1, 3-dipolar cycloaddition of azides and alkynes (CuAAC)-mediated nanosensors, as well as the functionalization of nanoprobes based on click chemistry. Nanoprobes including gold nanoparticles, quantum dots, magnetic nanoparticles and carbon nanomaterials are covered. We discuss the advantages of click chemistry-mediated nanosensors for biochemical assays, and give perspectives on the development of click chemistry-mediated approaches for clinical diagnosis and other biomedical applications.

Quantitative Detection of MicroRNA in One Step via Next Generation Magnetic Relaxation Switch Sensing

One-step, quantitative and rapid detection of microRNA (miRNA) in tumor cells or tissues can provide critical information for clinical diagnosis and cancer treatment. In this work, we develop a magnetic relaxation switch sensing (MRS)-based miRNA sensor using magnetic microparticle (1 μm in diameter, MM1000)-DNA probe-magnetic nanoparticle (30 nm in diameter, MN30) conjugates (MM1000-DNA-MN30). In the presence of target miRNA, DSN enzyme selectively cleaves the DNA tether after miRNA/DNA hybridization to release MN30 and leaves the miRNA intact to lead to the declustering of more MN30 than before. In contrast to conventional MRS by measuring the change of transverse relaxation time (ΔT2) induced by the aggregation or dissociation of magnetic particles in the presence of target, we use the cleaved MN30 from conjugates as the direct readout of ΔT2, which is more sensitive and stable. This MRS-based assay allows for one-step detection of 5 fM of miR-21 in urine samples, quantification of miR-21 from 100 cancer cells, and differentiation of the expression of miR-21 in tumor and surrounding tissues. The merits of this assay, rapidity, ability for quantitation, high sensitivity, and one-step operation, ensure a promising future in diagnostic technology.

Double-Enzymes-Mediated Bioluminescent Sensor for Quantitative and Ultrasensitive Point-of-Care Testing

We report an ultrasensitive, quantitative, and rapid bioluminescent immunosensor (ABS) for point-of-care testing (POCT) of the disease biomarker in clinical samples using double enzymes including alkaline phosphatase (ALP) and luciferase. In the presence of the biomarker, the ALP attached on the surface of immuno-nanocomplex dephosphorylates adenine triphosphate (ATP), subsequently inhibiting the ATP-luciferin-luciferase bioluminescent reaction. The highly sensitive response of ATP (picomolar level) allows for ultrasensitive detection of biomarker via the effective change of the bioluminescence intensity through ALP- and luciferase-catalyzed reactions, which can be quantitatively determined by a portable ATP detector. This ABS fulfills the criteria for POCT that performs sensitive (femtomolar level of biomarkers) and quantitative measurement quickly (less than 1 h) with minimal equipment (portable detector).

Point-of-Care Detection of β-Lactamase in Milk with a Universal Fluorogenic Probe

The illegal addition of β-lactamase (Bla) in milk to disguise β-lactam antibiotics has been a serious issue in the milk industry worldwide. Herein, we report a method for point-of-care detection of Bla based on a probe, Tokyo Green-tethered β-lactam (CDG-1), as a common substrate of various Blas (Bla A, B...) which can enzymatically convert CDG-1 (low fluorescence) to Tokyo Green (high fluorescence). This approach allows rapid screening of a broad spectrum of Blas in real milk samples within 15 min without any pretreatment. Combined with the immuno-magnetic separation, we achieved sensitive and quantitative detection of Bla (10(-5) U/mL), which provides a universal platform for screening and determining Blas in complex samples with high efficiency and accuracy.

Skiving stacked sheets of paper into test paper for rapid and multiplexed assay

Stacked paper skiving paves the way for industrial manufacturing of paper-based analytical devices for barcode assays. This paper shows that stacked sheets of paper preincubated with different biological reagents and skiving them into uniform test paper sheets allow mass manufacturing of multiplexed immunoassay devices and simultaneous detection of multiplex targets that can be read out by a barcode scanner. The thickness of one sheet of paper can form the width of a module for the barcode; when stacked, these sheets of paper can form a series of barcodes representing the targets, depending on the color contrast provided by a colored precipitate of an immunoassay. The uniform thickness of sheets of paper allows high-quality signal readout. The manufacturing method allows highly efficient fabrication of the materials and substrates for a straightforward assay of targets that range from drugs of abuse to biomarkers of blood-transmitted infections. In addition, as a novel alternative to the conventional point-of-care testing method, the paper-based barcode assay system can provide highly efficient, accurate, and objective diagnoses.

Bioorthogonal Reaction-Mediated ELISA Using Peroxide Test Strip as Signal Readout for Point-of-Care Testing

This work demonstrates a highly sensitive peroxide test strip (PTS)-based enzyme-linked immunosorbent assay (ELISA) for both qualitative and quantitative detection of drugs of abuse (morphine) and disease biomarkers (interleukin-6 and HIV-1 capsid antigen p24). This color-based PTS is a commercially available product with advantages of low cost, easy operation, and portability, and it is an ideal signal readout strategy in ELISA to simplify the immunoassay procedures and enable point-of-care testing (POCT). In addition, we introduce the bioorthogonal reaction that can effectively amplify the signal by controlling the cycles of bioorthogonal reaction to achieve the desirable sensitivity depending on different analytes. The limit of detection is 0.2 ng/mL for morphine, 3.98 pg/mL for interleukin-6, and 11.6 pg/mL for detection of HIV-capsid antigen (p24). This PTS-ELISA applies to both the qualitative and quantitative detection of IL-6 and p24 in clinical serum samples with good accuracy, which provides a promising tool for the POCT in clinical diagnosis.

Detection of Hepatitis B Virus M204I Mutation by Quantum Dot-Labeled DNA Probe

Quantum dots (QDs) are semiconductor nanoparticles with a diameter of less than 10 nm, which have been widely used as fluorescent probes in biochemical analysis and vivo imaging because of their excellent optical properties. Sensitive and convenient detection of hepatitis B virus (HBV) gene mutations is important in clinical diagnosis. Therefore, we developed a sensitive, low-cost and convenient QDs-mediated fluorescent method for the detection of HBV gene mutations in real serum samples from chronic hepatitis B (CHB) patients who had received lamivudine or telbivudine antiviral therapy. We also evaluated the efficiency of this method for the detection of drug-resistant mutations compared with direct sequencing. In CHB, HBV DNA from the serum samples of patients with poor response or virological breakthrough can be hybridized to probes containing the M204I mutation to visualize fluorescence under fluorescence microscopy, where fluorescence intensity is related to the virus load, in our method. At present, the limits of the method used to detect HBV genetic variations by fluorescence quantum dots is 103 IU/mL. These results show that QDs can be used as fluorescent probes to detect viral HBV DNA polymerase gene variation, and is a simple readout system without complex and expensive instruments, which provides an attractive platform for the detection of HBV M204I mutation.