Shape-memory microfluidic chips for fluid and droplet manipulation.

Fluid manipulation is an important foundation of microfluidic technology. Various methods and devices have been developed for fluid control, such as electrowetting-on-dielectric-based digital microfluidic platforms, microfluidic pumps, and pneumatic valves. These devices enable precise manipulation of small volumes of fluids. However, their complexity and high cost limit the commercialization and widespread adoption of microfluidic technology. Shape memory polymers as smart materials can adjust their shape in response to external stimuli. By integrating shape memory polymers into microfluidic chips, new possibilities for expanding the application areas of microfluidic technology emerge. These shape memory polymers can serve as actuators or regulators to drive or control fluid flow in microfluidic systems, offering innovative approaches for fluid manipulation. Due to their unique properties, shape memory polymers provide a new solution for the construction of intelligent and automated microfluidic systems. Shape memory microfluidic chips are expected to be one of the future directions in the development of microfluidic technology. This article offers a summary of recent research achievements in the field of shape memory microfluidic chips for fluid and droplet manipulation and provides insights into the future development direction of shape memory microfluidic devices.

Two-Dimensional Multi-parameter Cytometry Platform for Single-Cell Analysis.

A 2D flow cytometry platform, known as CytoLM Plus, was developed for multi-parameter single-cell analysis. Single particles or cells after hydrodynamic alignment in a microfluidic unit undergo first-dimension fluorescence and side scattering dual-channel optical detection. They were thereafter immediately directed to ICP-MS by connecting the microfluidic unit with a high-efficiency nebulizer to facilitate the second-dimension ICP-MS detection. Flow cytometry measurements of fluorescent microspheres evaluated the performance of CytoLM Plus for optical detection. 6434 fluorescence bursts were observed with a valid signal proportion as high as 99.7%. After signal unification and gating analysis, 6067 sets of single-particle signals were obtained with 6.6 and 6.2% deviations for fluorescence burst area and height, respectively. This is fairly comparable with that achieved by a commercial flow cytometer. Afterward, CytoLM Plus was evaluated by 2D flow cytometry measurement of Ag+-incubated and AO-stained MCF-7 cells. A program for 2D single-cell signal unification was developed based on the algorithm of screening in lag time window. In the present case, a lag time window of -4.2 ± 0.09 s was determined by cross-correlation analysis and two-parameter optimization, which efficiently unified the concurrent single-cell signals from fluorescence, side scattering, and ICP-MS. A total of 495 sets of concurrent 2D signals were screened out, and the statistical analysis of these single-cell signals ensured 2D multi-parameter single-cell analysis and data elucidation.

Open flow cytometer with the combination of 3D hydrodynamic single cell focusing and confocal laser-induced fluorescence detection.

Flow cytometry is among the most powerful tools for single-cell analysis, while the high cost and mechanical complexity of the commercial instrumentation limit the applications in personalized single-cell analysis. For this issue, we hereby construct an open and low-cost flow cytometer. It is highly compact to integrate the functions of (1) single cell aligning by a lab-made modularized 3D hydrodynamic focusing device, and (2) fluorescence detection of the single cells by a confocal laser-induced fluorescence (LIF) detector. The ceiling cost of the entire hardware for the LIF detection unit and 3D focusing device is $ 3200 and $ 400 respectively. A sheath flow velocity of 150 µL/min produces a focused sample stream of 17.6 µm × 14.6 µm at sample flow of 2 µL/min, based on the LIF response frequency and the laser beam spot diameter. The assay performance of the flow cytometer was evaluated by characterizing fluorescent microparticles and acridine orange (AO) stained HepG2 cells, producing throughputs of 40.5/s and 6.2/s respectively. Favorable assay precision and accuracy were demonstrated by the agreement of frequency histogram with imaging analysis, and good Gaussian-like distributions of fluorescent microparticles and AO-stained HepG2 cells. Practically, the flow cytometer was successfully applied for the evaluation of ROS generation in single HepG2 cells.

Open-channel microfluidic chip based on shape memory polymer for controllable liquid transport.

Open microfluidics has attracted increasing attention over the last decade because of its flexibility and simplicity with respect to cell culture and clinical diagnosis. However, traditional valves and pumps are difficult to integrate on open-channel microfluidic chips, in which a liquid is usually driven by capillary forces. Poor fluid control performance is a common drawback of open microfluidics. Herein, we proposed a method for controlling the liquid flow in open channels by controlling the continuous Laplace pressure induced by the deformation of the shape memory microstructures. The uniformly arranged cuboidal microcolumns in the open channels have magnetic/light dual responses, and the bending angle of the microcolumns can be controlled by adjusting Laplace pressure using near-infrared laser irradiation in a magnetic field. Laplace pressure and capillary force drove the liquid flow together, and the controllable fluid transport was realized by adjusting the hydrophilicity of the channel surface and the bending angle of the microcolumns. We demonstrated the controllability of the flow rate and the directional transport of water along a preset path. In addition, the start and stop of water transport were realized via local hydrophobic modification. The proposed strategy improves poor fluid control in traditional open systems and makes fluid flow highly controllable. We tried to extract and detect rhodamine B in tiny droplets on the open microfluidic chip, demonstrating the advantages of the proposed strategy in the separation and analysis of tiny samples.

Photothermal visual sensing of alkaline phosphatase based on the etching of Au@MnO2 core-shell nanoparticles.

Alkaline phosphatase (ALP), as a crucial enzyme involved in many physiological activities, is always used as one of the significant biomarkers in clinical diagnosis. Herein, a novel, simple, and effective photothermal quantitative method based on the etching of MnO2-coated gold nanoparticles (Au@MnO2 NPs) was established for ALP activity assay with a household thermometer-based visual readout. The photothermal effect of Au@MnO2 NPs is much higher than that of MnO2 NPs or Au NPs. The MnO2 shell of Au@MnO2 NPs can be etched by ascorbic acid, a product of ALP-catalyzed hydrolysis of 2-phospho-l-ascorbic acid. With the etching of Au@MnO2 NPs, the photothermal conversion efficiency decreased gradually, causing the decrease of the temperature increment of the solutions by degrees. A household thermometer, instead of large-scale and professional instruments, was used as a signal reader to realize the visual quantitative detection. The photothermal platform was used successfully for the determination of ALP with a wide linear range from 2.0 to 50 U/L and a detection limit as low as 0.75 U/L. Moreover, the inhibition efficiency of sodium vanadate for ALP activity was investigated, proving the photothermal quantitative method will be a potential platform for screening enzyme inhibitors. Such a sensitive, facile, and low-cost sensing assay provides a new prospect to develop platforms for point-of-care testing.

In-situ and amplification-free imaging of hERG ion channels at single-cell level using a unique core-molecule-shell-secondary antibody SERS nanoprobe.

Research on ion channels and their monoclonal antibodies plays a critical role in drug development and disease diagnosis. The current ion channel researches are often not conducted in the microenvironment for cells survival, which restricts the mechanism study of the links between the cell structure and the ion channel function. In this work, we synthesized gold core-4-mercaptobenzonitrile-sliver shell-goat anti-rabbit immunoglobulin G (Au@4-MBN@Ag@IgG) nanoparticles as surface-enhanced Raman scattering (SERS) nanoprobes for investigating the human ether-a-go-go related gene (hERG) potassium ion channel in cell membranes. This is the first attempt to study ion channels using SERS. Due to the unique core-molecule-shell structure and the silver shell of nanoprobes, strong and stable SERS signal was obtained. With the help of antibodies, the Au@4-MBN@Ag@IgG nanoprobes were captured by hERG antibodies and then bonded with hERG ion channels based on the sandwich immune response. The reporter molecule, 4-MBN, displayed a strong and sharp characteristic peak at 2233 cm-1 in the Raman silent region. The intensity of this peak denoted the concentration of antibodies and the expression of ion channel proteins. We successfully applied this amplification-free method for in-situ imaging the distribution of the hERG ion channel on the transfected HEK293 cell surface at the single-cell level. This hERG ion channel profiling strategy promises a maneuverable tool for ion channel research.

Tracking cellular transformation of As(III) in HepG2 cells by single-cell focusing/capillary electrophoresis coupled to ICP-MS.

The cellular metabolism of metals is highly critical to elucidate their potential cytotoxicity or cell protection mechanism. In this work, an asymmetric serpentine microfluidic device (ASMD) with high sampling efficiency and excellent focusing performance was developed for single-cell focusing. ASMD coupling with ICP-MS ensures single-cell assay to provide the information for trivalent arsenic (As(III)) uptake by HepG2 cells, which reveals the heterogeneity of cellular arsenic distribution, and elucidates the arsenic elimination behaviors in single HepG2 cells. Further, the metabolism and transformation of As(III) in HepG2 cells was tracked by hyphenating capillary electrophoresis (CE) separation with ICP-MS. The results for single-cell analysis and arsenic elimination kinetics illustrated that the half-life of arsenic elimination is 0.9 ± 0.04 h with the elimination constant of 0.77 ± 0.03, i.e., 77% of accumulated As in HepG2 cells may be eliminated per hour. Moreover, arsenobetaine (AsB) was identified to be the main metabolite and biotransformation species of As in HepG2 cells. ASMD-ICP-MS and CE-ICP-MS are powerful for tracking the fate of metals or metal drugs in single cells to comprehensively understand their metabolic pathway and transformation behaviors.

A nanohybrid of Prussian blue supported by boracic acid-modified g-C3N4 for Raman recognition of cell surface sialic acid and photothermal/photodynamic therapy.

Theranostic nanoplatforms with accurate diagnosis and effective therapy show a bright prospect for tumor treatments. Herein, a novel boracic acid-modified graphite carbon nitride and Prussian blue nanohybrid (PB@B-g-C3N4) was developed, which provides sialic acid-targeted Raman recognition and synergistic photothermal/photodynamic therapy in the near-infrared region. Owing to the specific interaction between boracic acid and sialic acid and Raman response at 2157 cm-1 of PB, the nanohybrids exhibit high specificity and Raman sensitivity for detection of the overexpressed sialic acid on tumor cells. Moreover, the photothermal conversion efficiency of PB@B-g-C3N4 is as high as 47.0% with 808 nm laser irradiation due to the enhanced absorbance of PB@B-g-C3N4. PB@B-g-C3N4 also possesses excellent photodynamic activity, which is attributed to the energy transfer of PB (type I) and electron transfer between PB and B-g-C3N4 (type II). This nanotheranostic agent for Raman recognition of cancer markers and synergistic photothermal/photodynamic therapy holds great potential for the development of efficient theranostic nanoplatforms.

In situ monitoring PUVA therapy by using a cell-array chip-based SERS platform.

Psoralen ultraviolet A (PUVA) therapy has thrived as a promising treatment for psoriasis. However, overdose of PUVA treatment will cause side-effects, such as melanoma formation. And these side-effects are often ignored during PUVA therapy. Hence, in situ monitoring therapeutic response of PUVA therapy is important to minimize side-effects. Aberrant expression of tyrosinase (TYR) has been proved to be associated with melanoma, indicating that TYR is a potential target for evaluation of PUVA therapy. Herein, we reported a strategy for in situ monitoring TYR activity during PUVA therapy by using a cell-array chip-based SERS platform. The cell-array chip was used to simulate cell survival environment for cell culture. Capture of single cells and living cell analysis were realized in the isolated microchambers. An enzyme-induced core-shell self-assembly substrate was used to evaluate TYR activity in living cells during PUVA therapy. The gold nanoparticle modified with a SERS reporter, 4-mercaptobenzonitrile (4-MBN), was used as the core. In the presence of oxygen and TYR, hydroxylation of l-tyrosine occurred, leading to the reduction of silver ion on the surface of gold cores. The growth of silver shells was accompanied by the increased SERS intensity of the reporter, which is related directly to TYR activity. The detection limit for TYR activity is 0.45 U/mL. Upregulation of TYR activity was successfully monitored after PUVA therapy. Notably, real-time and in situ information of therapeutic response can be obtained through monitoring PUVA therapy by using a cell-array chip-based SERS platform, which has great potential to guide the clinical application of PUVA therapy.

Two-Dimensional Cytometry Platform for Single-Particle/Cell Analysis with Laser-Induced Fluorescence and ICP-MS.

A two-dimensional cytometry platform (CytoLM) with high sensitivity and high temporal resolution is developed for single-particle and single-cell sampling and analysis. First, a Dean flow-assisted vortex capillary cell sampling (VCCS) unit confines the sample stream in curved flow and drives to focus and align the particles or cells in a small probe volume. By coupling VCCS to a laser-induced fluorescence (LIF) detector with data acquisition and processing capability, a high-throughput single-particle/cell analysis system (VCCS-LIF) was established. The particle analysis throughput of 119.42/s and a detection recovery of 78.20 ± 1.75% were achieved at a density of 9.16 × 104/mL for fluorescent particles, and the cell analysis throughput is 48.20/s at a density of 1.5 × 105/mL. Second, the CytoLM platform is constructed by hyphenating VCCS-LIF with inductively coupled plasma mass spectrometry (ICP-MS). In the analysis of HepG2 cells by Ag+ incubation and AO staining, 10,760 fluorescence bursts and 3068 MS events were observed in 240 s. Invalid signals due to undispersed cells were controlled at 3.80% for LIF and 1.01% for MS, with a proportion of effective signal of >96.20%. After peak identification and integral processing of the original data, the statistical results including peak area, height, width, and spacing are obtained concurrently and the information on concentration and elemental quantification of single cells is evaluated. CytoLM facilitates high-throughput, multi-dimensional, and multi-parameter characterization of particles and cells, and it may provide vast potential in life science analysis.

Alkaline phosphatase-regulated in situ formation of chromogenic probes for multicolor visual sensing of biomarkers.

Alkaline phosphatase (ALP), as an immunological label, is widely used in biochemical assays. Here, a simple yet effective strategy for ALP activity detection was proposed on the basis of in situ formation of Prussian blue nanoparticles and polychromatic superposition effect. Firstly, ascorbic acid, a product from ALP-catalyzed hydrolysis of 2-phospho-l-ascorbic acid (AAP), converted yellow ferricyanide into ferrocyanide. Then, the specific reaction between ferrocyanide and ferric ions (Fe3+) initiated the generation of Prussian blue nanoparticles in situ. Meanwhile, the residual AAP chelated with Fe3+, and a stable Fe3+-AAP complex was quickly formed. When Prussian blue nanoparticles mixed with brown Fe3+-AAP complex and yellow ferricyanide at different ratios, a distinct color variation was presented. Therefore, a sensitive multicolor assay of ALP activity with a detection limit of 1.0 U/L was realized by simply blending commercially available reagents. Furthermore, magnetic sandwich and competitive sensing platforms for multiple biomarkers detection were constructed by combining the ALP-regulated multicolor system with the well-developed aptasensor. The feasibility of the sensors was convincingly demonstrated by using thrombin and prostate specific antigen as model targets. In addition, the proposed multicolor strategy was employed for evaluating inhibition efficiency, and shows potential in visual screening of enzyme inhibitors. Such a facile, sensitive and low-cost sensing strategy provides a new perspective to develop universal platforms of point-of-care testing.

A digital microfluidic platform based on a near-infrared light-responsive shape-memory micropillar array.

In this work, we developed a digital microfluidic platform based on a shape memory micropillar array responsive to near-infrared light, and the droplets were programmatically manipulated through light-induced micropillar deformation. The micropillar array was constructed on the surface of a poly(ethylene-vinyl acetate) copolymer, a shape memory polymer sensitive to near-infrared light. Before droplet manipulation, the micropillar array was kept temporarily tilted by heating and pressing. Under the irradiation of a near-infrared laser, the micropillar array achieved the transition from the temporary shape to the original shape. Temperature gradient and micropillar deformation caused by near-infrared light irradiation produce the driving force for droplet movement. The movement of the laser mounted on an electronically controlled displacement platform was controlled by a computer to achieve the programmed control of the droplets. Moreover, we demonstrated light-manipulated droplet movement and fusion, and achieved ascorbic acid detection using this digital microfluidic platform. In particular, the micropillar array chip is able to manipulate droplets in a wide range of 0.1 µL to 10 µL. The proposed digital microfluidic platform will broaden the application of digital microfluidic technology in analytical chemistry and biomedicine.

Enhanced chemodynamic therapy at weak acidic pH based on g-C3N4-supported hemin/Au nanoplatform and cell apoptosis monitoring during treatment.

Chemodynamic therapy (CDT), inducing tumor cell apoptosis through Fenton reaction to produce hydroxyl radical (·OH), is an emerging cancer treatment technology. Highly efficient Fenton catalytic reactions usually take place at a low pH environment. Utilizing graphitic carbon nitride supported hemin and Au nanoparticles (g-C3N4/hemin/Au) as a novel biomimetic nanocatalyst, we achieve an enhanced CDT for inducing tumor cell apoptosis in the presence of excess H2O2, and reveal the molecular events during the CDT-induced apoptosis. The prepared g-C3N4/hemin/Au nanohybrids exhibit excellent Fenton catalytic activity for the generation of highly toxic ·OH at weak acidic and neutral condition, which breaks through the limitation of traditional acidity-dependent response. The Fenton catalytic mechanism was also studied. The Fenton efficiency is primarily enhanced by the high affinity between nanohybrids and H2O2, and the transformation of Fe(III) to Fe(IV)=O without the formation of iron hydrate precipitation. Moreover, the intracellular molecular events during the CDT process were monitored. Phenylalanine metabolism was perturbed with protein degradation and DNA structures were damaged, which eventually lead to cell apoptosis. This study provides a significant guidance for the further development of more effective CDT platforms.

Visual detection of acid phosphatase based on hollow mesoporous manganese dioxide nanospheres.

Acid phosphatase is widely used as a clinical indicator because of its close correlation with a variety of diseases. Herein, a label-free and colorimetric sensing method for detecting the activity of acid phosphatase was constructed based on hollow mesoporous manganese dioxide nanospheres. The nanospheres exhibit superior oxidase-like property and can oxidize colorless 3,3',5,5'-tetramethylbenzidine (TMB) to yellow TMB2+. Ascorbic acid from acid phosphatase-catalyzed hydrolysis of L-ascorbic acid-2-phosphate will inhibit the oxidization reaction, igniting vivid color variation. On the basis of this obvious multicolor change, the visual detection of acid phosphatase was achieved. Compared with the single-color change, the multicolor colorimetric method is more conducive for naked-eye discrimination. The absorbance difference at 450 nm exhibits a linear relationship with the concentration of acid phosphatase ranging from 1.0 to 25 U L-1, with a detection limit as low as 0.45 U L-1. Acid phosphatase in human serum samples was successfully determined. Moreover, the inhibition efficiency of NaF for acid phosphatase activity was investigated, proving the proposed colorimetric method will be a potential platform for screening acid phosphatase inhibitors and discovering new drugs.

Inertial-Force-Assisted, High-Throughput, Droplet-Free, Single-Cell Sampling Coupled with ICP-MS for Real-Time Cell Analysis.

Single-cell analysis facilitates perception into the most essential processes in life's mysteries. While it is highly challenging to quantify them at the single-cell level, where precise single-cell sampling is the prerequisite. Herein, a real-time single-cell quantitative platform was established for high-throughput droplet-free single-cell sampling into time-resolved (TRA) ICP-MS and real-time quantification of intracellular target elements. The concentrated cells (2 × 106 cells mL-1) were spontaneously and orderly aligned in a spiral microchannel with 104 periodic dimensional confined micropillars. The quantification is conducted simultaneously by internal standard inducing from another branch channel in the chip. The flow-rate-independent feature of single-cell focusing into an aligned stream within a wide range of fluidic velocities (100-800 µL min-1) facilitates high-throughput, oil-free, single-cell introduction into TRA-ICP-MS. The system was used for real-time exploration of intracellular antagonism of Cu2+ against Cd2+. an obvious antagonistic effect was observed for the MCF-7 cell by culturing for 3, 6, 9, and 12 h with 100 µg L-1 Cd2+ and 100 µg L-1 Cu2+, and a rivalry rate of 12.8% was achieved at 12 h. At identical experimental conditions, however, limited antagonistic effect was encountered for a bEnd3 cell within the same incubation time period, with a rivalry rate of 4.81%. On the contrary, an antagonistic effect was not observed for the HepG2 cell by culturing for 6 h, while an obvious antagonistic effect was found by further culturing to 12 h, with a rivalry rate of 10.43%. For all three cell lines, significant heterogeneity was observed among individual cells.

Colloidal photonic crystal array chip based on nanoparticle self-assembly on patterned hydrophobic surface for signal-enhanced fluorescent assay of adenosine.

A controllable approach for preparing a portable colloidal photonic crystal (CPC) array chip is presented. The approach was inspired by the confinement effect of nanoparticle self-assembly on patterned surface. Hydrophobic polydimethylsiloxane substrate with reproducible micro-region array was fabricated by soft-lithography. The substrate was employed as the patterned template for self-assembly of monodisperse polystyrene nanoparticles. The CPC units can be prepared in several minutes, and exhibit consistent reflection wavelength. By adjusting the size of polystyrene nanoparticles and the shape of micro-regions, CPC units with multiple structure, colors and geometries were obtained. The CPC array chip features fluorescence enhancement owing to the optical modulation capability of the periodic nanostructure of the self-assembled CPC. With the reflection wavelength (523 nm) of green CPC units overlapping the emission wavelength (520 nm, with excitation wavelength of 490 nm) of 6-carboxyfluorescein-labeled DNA probe, the fluorescence intensity increased more than 10-fold. For signal-amplified assay of adenosine, the concentration range of linear response was 5.0 × 10-5 mol L-1 to 1.0 × 10-3 mol L-1, and the limit of detection was 1.3 × 10-6 mol L-1. Because of the enhancement effect of photonic crystal, the fluorescence images were more readable from the CPC array chip, compared with those from the planar substrate. The chip has potential applications in multiplex determination with high-throughput via encoding strategy based on the tunable structure, color or geometric shape. Graphical abstractSchematic diagram of signal-enhanced fluorescent detection of adenosine based on the colloidal photonic crystal array chip (PDMS, polydimethylsiloxane; PS NPs, polystyrene nanoparticles; CPC, colloidal photonic crystal; GO, graphene oxide; FAM, 6-carboxyfluorescein).

Controlled formation of porous CuCo2O4 nanorods with enhanced oxidase and catalase catalytic activities using bimetal-organic frameworks as templates.

Porous structures with highly dispersed and active catalytic sites are vital to improve the catalytic activity and stability of artificial enzyme-related catalytic reactions. Herein, a novel nanorod-like bimetal-organic framework serving as porous support and supplier of Co2+ and Cu2+ was used to prepare a beneficial porous metal oxide. By optimizing the calcination temperature, the composition of calcined product can be controlled and the nanorods with isolated and highly active CuCo2O4 nanoparticles were obtained. The porous CuCo2O4 nanorods exhibit a pH-dependent catalytic property, that is, they behave as oxidase in acid conditions and catalase in alkaline conditions. The CuCo2O4 nanorods perform dual-enzyme catalytic activity superior to monometallic oxides. What's more, compared with the reported Co3O4 nanoparticles, Co3O4/CuO hollow nanocage hybrids and NiCo2O4 mesoporous spheres, the porous CuCo2O4 nanorods show higher affinity to 3,3',5,5'-tetramethylbenzidine with a lower Km value. The superior dual-enzyme catalytic activities of CuCo2O4 nanorods benefit from the high catalytic activity of binary metal oxides and structural stability. After incubating in a wide range of pHs, temperatures and ionic strengths, the catalytic activity of CuCo2O4 nanorods can be maintained. The oxidase activity of CuCo2O4 nanorods can be inhibited in the presence of ascorbic acid, which can be applied in effective detection of ascorbic acid. This study opens a new path to prepare stable and highly active porous artificial enzymes.

Accurate quantitative detection of cell surface sialic acids with a background-free SERS probe.

Sialic acid (SA) is a special monosaccharide widely distributed at the termini of sugar chains on the cell surface, and its expression level is closely connected with various biological and pathological processes. Therefore, accurate quantitative detection of SA on cancer cell surface is of great significance for clinical diagnosis and therapy. Here, we developed a whole-surface accessible method of accurate SERS quantification of SA level on a single cell, in which silver nanoparticles functionalized with 4-mercaptophenylboric acid and 4-mercaptobenzenitrile was used as the background-free SERS probe. The cyano group on the nanoprobe showed a unique Raman shift at 2232 cm-1, where most of the biological samples have no Raman response. Meanwhile, the boronic acid group had high specificity to SA molecules at physiological pH. The expression level of SA can be accurately quantitated on the basis of the CN Raman signal. The average number of expressed SA molecules on the surface of a single HeLa cell was 4.6 × 107. And SERS imaging of a single cell was achieved at 2232 cm-1 without biological interference. We evaluated SA expression level on the surface of different cancer cells and dynamically monitored SA expression under the influence of drugs. The proposed approach is accurate as well as sensitive for background-free quantification of SA on cell surface, which is promising for revealing the relationship between tumors and cell surface glycosylation.

Multiplexed detection of micro-RNAs based on microfluidic multi-color fluorescence droplets.

In this work, simple, rapid, and low-cost multiplexed detection of tumor-related micro-RNAs (miRNAs) was achieved based on multi-color fluorescence on a microfluidic droplet chip, which simplified the complexity of light path to a half. A four-T-junction structure was fabricated to form uniform nano-volume droplet arrays with customized contents. Multi-color quantum dots (QDs) used as the fluorescence labels were encapsulated into droplets to develop the multi-path fluorescence detection module. We designed an integrated multiplex fluorescence resonance energy transfer system assisted by multiple QDs (four colors) and one quencher to detect four tumor-related miRNAs (miRNA-20a, miRNA-21, miRNA-155, and miRNA-221). The qualitative analysis of miRNAs was realized by the color identification of QDs, while the quantitative detection of miRNAs was achieved based on the linear relationship between the quenching efficiency of QDs and the concentration of miRNAs. The practicability of the multiplex detection device was further confirmed by detecting four tumor-related miRNAs in real human serum samples. The detection limits of four miRNAs ranged from 35 to 39 pmol/L was achieved without any target amplification. And the linear range was from 0.1 nmol/L to 1 µmol/L using 10 nL detection volume (one droplet) under the detection speed of 320 droplets per minute. The multiple detection system for miRNAs is simple, fast, and low-cost and will be a powerful platform for clinical diagnostic analysis. Graphical abstract.

High sensitivity and non-background SERS detection of endogenous hydrogen sulfide in living cells using core-shell nanoparticles.

Endogenous hydrogen sulfide (H2S) exists in multiple physiological processes. In order to further understand the action mechanism of H2S in cells and human body, we proposed a smart surface-enhanced Raman scattering (SERS) nanoprobe, Au core-4-mercaptobenzonitrile-Ag shell nanoparticle (Au@4-MBN@Ag), for the detection of endogenous H2S in living cells based on the reaction between Ag shell and sulfide species. 4-MBN was selected as the SERS reporter to avoid interference from cellular molecules. With the sulfide concentration increasing, the Ag2S constantly formed, and consequently the SERS signal intensity of Au@4-MBN@Ag gradually decreased owing to the weaker SERS activity of Ag2S. With the nanoprobes, this method not only offers a high sensitivity for H2S detection at an nM level, but also achieves the goal of non-background analysis. It displays satisfactory anti-interference capability and a good linear relationship with sulfide concentration ranging from 50 nM to 500 µM, and an estimated detection limit is 0.14 nM. The Au@4-MBN@Ag nanoprobes were successfully applied to detect endogenous H2S in living HepG2 cells stimulated by pyridoxal 5-phosphate monohydrate. This work offers a potential analytical method in the related research of H2S physiological function.