Sifeng Mao

Inkjet Nanoinjection for High-Thoughput Chemiluminescence Immunoassay on Multicapillary Glass Plate

We report a novel chemiluminescence diagnosis system for high-throughput human IgA detection by inkjet nanoinjection on a multicapillary glass plate. As proof-of-concept, microhole-based polydimethylsiloxane (PDMS) sheets were aligned on a multicapillary glass plate to form a microwell array as microreactors for enzyme-linked immunosorbent assay (ELISA). The multicapillary glass plate was utilized as a switch that controlled the holding/passing of the solution. Further, anti-IgA-labeled polystyrene (PS) microbeads was assembled into the microwell array, and an inkjet nanoinjection was specially used to distribute the sample and reagent solution for chemiluminescence ELISA, enabling high-throughput detection of human IgA. As a result, the performance of human IgA tests revealed a wider range for the calibration curve and a lower limit of detection (LOD) of 0.1 ng mL(-1) than the ELISA by a standard 96-well plate. The analysis time and reagent consumption were significantly decreased. The IgA concentrations in saliva samples were determined after 10000-fold dilution by the developed ELISA system showing comparable results by conventional immune assay with 96-wells. Thus, we believe that the inkjet nanoinjection for high-throughput chemiluminescence immunoassay on a multicapillary glass plate will be promising in disease diagnosis.

Cell signaling analysis by mass spectrometry under coculture conditions on an integrated microfluidic device.

A microfluidic device was integrated in a controlled coculture system, in which the secreted proteins were qualitatively and semiquantitatively determined by a directly coupled mass spectrometer. PC12 cells and GH3 cells were cocultured under various conditions as a model of the regulation of the organism by the nervous system. A micro-solid phase extraction (SPE) column was integrated in order to remove salts from the cells secretion prior to mass spectrometry detection. A three layer polydimethylsiloxane (PDMS) microfluidic device was fabricated to integrate valves for avoiding contamination between the cells coculture zone and the pretreatment zone. Electrospray ionization (ESI)-quadrupole (Q)-time of flight (TOF)-mass spectrometry was employed to realize highly sensitive qualitative analysis and to implement semiquantitative analysis. Furthermore, cell migrations under various coculture conditions were observed and discussed. The inhibition on growth hormone secretion from GH3 cells by dopamine released from PC12 cells was investigated and demonstrated. Thus, the developed platform provides a useful tool on cell to cell signaling studies for disease monitoring and drug delivery control.

In Situ Scatheless Cell Detachment Reveals Connections Between Adhesion Strength and Viability at Single-Cell Resolution

Single-cell biology provides insights into some of the most fundamental processes in biology and promotes the understanding of lifes mysteries. As the technologies to study single-cells expand, they will require sophisticated analytical tools to make sense of various behaviors and components of single-cells as well as their relations in the adherent tissue culture. In this paper, we revealed cell heterogeneity and uncovered the connections between cell adhesion strength and cell viability at single-cell resolution by extracting single adherent cells of interest from a standard tissue culture by using a microfluidic chip-based live single-cell extractor (LSCE). We believe that this method will provide a valuable new tool for single-cell biology.

Generation of controlled monodisperse porous polymer particles by dipped inkjet injection

A straightforward approach for the preparation of monodisperse porous polymer particles by inkjet technology is reported. Uniform droplets of an aqueous polymer solution were ejected from an inkjet, the nozzle of which was immersed in an organic phase. The subsequent dehydration of the droplets by solvent extraction resulted in the formation of porous polymer microspheres. Particles with a narrow size distribution could be obtained using the present system in a single step. Parameters for polymer droplet generation in an organic phase, including polymer properties, organic solvent, and inkjet waveform, were investigated. The relationship between the final particle diameter and the initial polymer droplet size were also examined. The diameters of the particles could be precisely controlled by changing the waveform exerted on the inkjet, thus producing particles with diameters in the range of 15 μm–60 μm with a coefficient of variation (CV) in the range of 2.7–4.8%. It was also possible to produce hollow porous polymer particles by simply decreasing the concentration of the polymer solution. The results demonstrate the merit of the inkjet-based method for the generation of monodisperse porous polymer particles. The method has the potential for producing functional polymer particles via the incorporation of functional or smart materials.

Online monodisperse droplets based liquid–liquid extraction on a continuously flowing system by using microfluidic devices

In this work, we developed a chip-based continuously flowing system for online liquid–liquid extraction by using monodisperse droplets, and the droplets containing target compounds after extraction were collected on the microchip. Monodisperse droplets of extracting reagent were on-chip generated and dispersed in continuously flowing sample solution, and then liquid–liquid extraction happened when organic droplets moved forward with sample solution flow. After liquid–liquid extraction, guiding tracks were designed in our system to capture the organic droplets, which allowed the aqueous solution flowing over at the same time. Structures of the microchip were optimized to achieve generation and collection of monodisperse droplets more efficiently. To verify the feasibility of droplet based liquid–liquid extraction on the established platform, we used aqueous solution containing 10 μM fluorescein sodium as sample solution and butanol as the extracting reagent to perform the whole procedures. Fluorescein sodium was successfully extracted by the butanol droplets, and the butanol droplets were successfully collected. The results demonstrated that the developed microfluidic device was a useful tool for monodisperse droplet based liquid–liquid extraction. As all the procedures including droplet generation, extraction and collection were performed on one chip, the established platform had the potential of time-resolved monitoring.

A reversibly electro-controllable polymer brush for electro-switchable friction

We report electrically switchable friction, which is reversibly manipulated by an external electric field via an electro-controllable polymer branched brush. The electro-controllable polymer branched brush, poly-sodium allyloxy hydroxypropyl sulfonate (poly-AHPS), was grafted onto an electrically conductive indium tin oxide (ITO) glass, and it demonstrated the capability of switching states by responding to the external electric field in a controllable manner. The two states of poly-AHPS branched brushes – stretched polymer brush arrays when negatively charged and rolled polymer knolls when positively charged – correspond to two different friction factors. Meanwhile, the shifting span for friction factors would be compressed by sandwiching a polar molecule medium or a nonpolar molecule medium, and exhibited a medium-controllable character. The proposed electric-field-responsive polymer brush is expected to be a manoeuvrable matrix for automatic control of friction, and to find wide application in automatization, intelligent manipulation and actuators.

The use of an inkjet injection technique in immunoassays by quantitative on-line electrophoretically mediated microanalysis

The paper describes a new online quantitative electrophoretically mediated microanalysis (EMMA) for use in immunoassays based on the unique drop-by-drop introduction of a sample by means of an inkjet for capillary electrophoresis (CE). Plugs of a fluorescein-labeled antibody (Anti-humanIgG-DyL550) and human IgG were alternately injected into a capillary using the inkjet, followed by the merging of the plugs and the subsequent immune reaction. The antigen-antibody complex that was formed in the merged zone was then separated by CE. As a proof-of-concept, the method was used to determine human IgG. As a result, both the consumption of the reaction solution and the analysis time were significantly reduced. The method showed a wide linear range (10-2000ngmL-1, R2=0.9912) of calibration and the detection limit (5ngmL-1) was substantially lower than that by for conventional methods.

Combination of nano-material enrichment and dead-end filtration for uniform and rapid sample preparation in matrix-assisted laser desorption/ionization mass spectrometry

Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is a fast analysis tool for the detection of a wide range of analytes. However, heterogeneous distribution of matrix/analyte cocrystal, variation in signal intensity and poor experimental reproducibility at different locations of the same spot means difficulty in quantitative analysis. In this work, carbon nanotubes (CNTs) were employed as adsorbent for analyte cum matrix on a conductive porous membrane as a novel mass target plate. The sample pretreatment step was achieved by enrichment and dead-end filtration and dried by a solid-liquid separation. This approach enables the homogeneous distribution of analyte in the matrix, good shot-to-shot reproducibility in signals and quantitative detection of peptide and protein at different concentrations with correlation coefficient (R2) of 0.9920 and 0.9909, respectively. The simple preparation of sample in a short time, uniform distribution of analyte, easy quantitative detection, and high reproducibility makes this technique useful and may diversify the application of MALDI-MS for quantitative detection of a variety of proteins.

A dual-functional microfluidic chip for on-line detection of interleukin-8 based on rolling circle amplification

Interleukin 8 (IL-8), also known as C-X-C motif ligand 8(CXCL8), is a proinflammatory chemokine functioned in neutrophil chemotaxis and activation. And it plays an important role in the process of glioma stem-like cell vascularization in the latest research. Herein, a dual-function microfluidic biosensor based on rolling circle amplification (RCA) was fabricated for cell culture and online IL-8 detection. A microfluidic chip was designed with two high passages connected by the vertical channels. One of the channels with immobilized capture antibody was prepared for IL-8 detection and another channel for cell culture. Immunoassays were achieved by a sandwich structure consisting of antibodies, IL-8, and aptamers. Signal amplification was mainly due to RCA and biotin-streptavidin linkage. The linear range for IL-8 was 7.5 -120pgmL-1 in this assay. Moreover, the developed method was successfully applied to detect the IL-8 in tumor-derived endothelial cells (TDEC) and Human Umbilical Vein Endothelial cells (HUVEC) under chemical hypoxia condition. Semi-quantitative detection of IL-8 consumption in HUVEC cells in low oxygen condition was also achieved. These results were in statistical agreement with those obtained by commercial assay of enzyme-linked immunoassay kit (ELISA). The microfluidic chip based biosensor reported hereby has a large prospect in the basic research and clinical diagnosis of cancer stem cell.

Shear Stress-Enhanced Internalization of Cell Membrane Proteins Indicated by a Hairpin-Type DNA Probe

Shear stress is an important mechanical stimulus that plays a critical role in modulating cell functions. In this study, we investigated the regulating effects of shear stress on the internalization of cell membrane proteins in a microfluidic chip. A hairpin-type DNA probe was developed and indiscriminately anchored to the cell surface, acting as an indicator for the membrane proteins. When cells were exposed to shear stress generated from fluid cell medium containing external proteins, strong fluorescence was emanated from intracellular regions. With intensive investigation, results revealed that shear stress could enhance the specific cell endocytosis pathway and promote membrane protein internalization. This process was indicated by the enhanced intracellular fluorescence, generated from the internalized and mitochondria accumulated DNA probes. This study not only uncovered new cellular mechanotransduction mechanisms but also provided a versatile method that enabled in situ and dynamic indication of cell responses to mechanical stimuli.