Qiushui Chen

Qualitative and Quantitative Analysis of Tumor Cell Metabolism via Stable Isotope Labeling Assisted Microfluidic Chip Electrospray Ionization Mass Spectrometry

In this work, a stable isotope labeling assisted microfluidic chip electrospray ionization mass spectrometry (SIL-chip-ESI-MS) platform for qualitative and quantitative analysis of cell metabolism was developed. Microfluidic cell culture, drug-induced cell apoptosis analysis, and cell metabolism measurements were performed simultaneously on the specifically designed device. MCF-7 cells were cultivated in vitro and exposed in anticancer agent (genistein and genistein-d(2)) for cell-based drug assay. A dual-isotopic labeling was presented for effective qualitative analysis of multiplex metabolites. Interestingly, three coeluting pairs of isotopomers appeared with an m/z difference of two. Despite complex biological matrixes, they can be easily recognized and identified by chip-ESI-MS/MS, which significantly facilitates candidate biomarker discovery. The quantitative performance of this system was evaluated using genistein as a model drug by means of stable isotope dilution analysis. The linear equation obtained is y = 0.06x - 3.38 × 10(-3) (R(2) = 0.995) at the dynamic range from 0.5 to 40 μM. The detection limit is 0.2 μM. The method shows an excellent stability of 2.2% relative standard deviation (RSD) and a good repeatability of 5.5% RSD. Our results have successfully demonstrated the capability of selective and quantitative analysis of cell-based drug absorption and metabolites with high stability, sensitivity, and repeatability on the chip-ESI-MS system. Consequently, the present device shows promise as a high-throughput, low-cost, and online platform for cell metabolism studies and drug screening processes.

Microfluidic isolation of highly pure embryonic stem cells using feeder-separated co-culture system

Engineered artificial tissues from stem cells show great potential in regenerative medicine, disease therapies and organ transplantation. To date, stem cells are typically co-cultured with inactivated feeder layers to maintain their undifferentiated state, and to ensure reliable cell purity. Herein, we propose a novel microfabricated approach for feeder-separated coculture of mouse embryonic stem (mES) cells on polydimethylsiloxane (PDMS) porous membrane-assembled 3D-microdevice. Normal mouse embryonic fibroblasts (mEFs) without inactivation were specifically co-cultured with mES cells, resulting in the formation of mES cell colonies on spatially controlled co-culture with feeder layers. An excellent undifferentiated state was confirmed by the expressions of Nanog, octamer binding protein 4 (Oct-4) and alkaline phosphatase (ALP) after 5 days culture. As a result, with the significant advantages of efficiency and simplicity, pure mES cell populations (a purity of 89.2%) from mEFs co-cultures were easily collected without any further purification or separation.

Oxygen-induced cell migration and on-line monitoring biomarkers modulation of cervical cancers on a microfluidic system.

In this work, we report an integrated microfluidic device for cell co-culture under different concentrations of oxygen, in which the secreted protein VEGF165 was on-line qualitatively and semi-quantitatively analyzed by functional nucleic acid, hemin, ABTS and peroxide system. This microfluidic platform allowed investigation of various oxygen and distances effect on cell-to-cell communication. Besides, the microfluidic device was used for real-time analysis of VEGF165 protein by aptamer-functionalized microchannels. Under 5% O2 condition, we found that the migration of CaSki cells was faster than the migration of human umbilical vein endothelial cells. However, the migration of CaSki cells was slower than the migration of HUVECs under 15% O2 condition. Moreover, the shorter intercellular distances, the quicker cells migration. Furthermore, HIF-1α and VEGF165 genes, ROS were analyzed, and the results would provide new perspectives for the diagnosis and medical treatment of cervical cancer.

Assay of multiplex proteins from cell metabolism based on tunable aptamer and microchip electrophoresis

A simple and rapid method for multiplex protein assay based on tunable aptamer by microchip electrophoresis has been developed. Different lengths of aptamers can modulate the electrophoretic mobility of proteins, allowing the protein molecules to be effectively separated in hydroxyethyl cellulose buffer with 1.00 mM magnesium ion. A non-specific DNA was exploited as an internal standard to achieve the quantitative assay and to reduce the interference. A fluorescence dye SYBR gold was exploited to improve the sensitivity and to suppress the interference from sample matrix. Under optimum conditions, quantitative assay of PDGF-BB (R(2)=0.9986), VEGF165 (R(2)=0.9909), and thrombin (R(2)=0.9947) were achieved with a dynamic range in the 5.00-150.0 nM and RSDs in the 5.87-16.3% range. The recoveries were varied from 83.6% to 113.1%. Finally, the proposed method was successfully applied to analyze cell secretions, and then the concentration of PDGF-BB and VEGF165 were detected from 5.15 nM to 2.03 nM, and 3.14 to 2.53 nM, respectively, indicating the established method can be used to analyze cell secretions.

Engineering Cell-Compatible Paper Chips for Cell Culturing, Drug Screening, and Mass Spectrometric Sensing

Paper-supported cell culture is an unprecedented development for advanced bioassays. This study reports a strategy for in vitro engineering of cell-compatible paper chips that allow for adherent cell culture, quantitative assessment of drug efficiency, and label-free sensing of intracellular molecules via paper spray mass spectrometry. The polycarbonate paper is employed as an excellent alternative bioscaffold for cell distribution, adhesion, and growth, as well as allowing for fluorescence imaging without light scattering. The cell-cultured paper chips are thus amenable to fabricate 3D tissue construction and cocultures by flexible deformation, stacks and assembly by layers of cells. As a result, the successful development of cell-compatible paper chips subsequently offers a uniquely flexible approach for in situ sensing of live cell components by paper spray mass spectrometry, allowing profiling the cellular lipids and quantitative measurement of drug metabolism with minimum sample pretreatment. Consequently, the developed paper chips for adherent cell culture are inexpensive for one-time use, compatible with high throughputs, and amenable to label-free and rapid analysis.

Statistical single-cell analysis of cell cycle-dependent quantum dot cytotoxicity and cellular uptake using a microfluidic system

This paper reports a novel method for the statistical analysis of quantum dot (QD) cytotoxicity and cellular uptake based on single cell cycles, which is part of a series of works on the study of QD cytotoxicity using a microfluidic system (Lab Chip, 2012, 12, 3474–3480; 2013, 13, 1948–1954). The specially designed microfluidic system consisted of a polydimethylsiloxane (PDMS) microwell array for single-cell arrangement and microchannels for QD solution diffusion, enabling effective control of stable cell density and the interdistance between them, as well as maintaining a constant QD concentration with no disturbance of the fluids which can affect cellular uptake. We showed that the treatment of QDs had no influence on cell cycles. However, the QD cytotoxicity was found to be dependent on cellular uptake in various cell cycle phases, because the accumulation and dilution of QDs happened in single cell cycles. The rank of QD cytotoxicity was G2/M > S > G0/G1. Thus, this technology could serve as a new strategy to investigate otherwise inaccessible mechanisms governing nanoparticle cytotoxicity.

Flexible control of cellular encapsulation, permeability, and release in a droplet-templated bifunctional copolymer scaffold.

Designing cell-compatible, bio-degradable, and stimuli-responsive hydrogels is very important for biomedical applications in cellular delivery and micro-scale tissue engineering. Here, we report achieving flexible control of cellular microencapsulation, permeability, and release by rationally designing a diblock copolymer, alginate-conjugated poly(N-isopropylacrylamide) (Alg-co-PNiPAM). We use the microfluidic technique to fabricate the bifunctional copolymers into thousands of mono-disperse droplet-templated hydrogel microparticles for controlled encapsulation and triggered release of mammalian cells. In particular, the grafting PNiPAM groups in the synthetic cell-laden microgels produce lots of nano-aggregates into hydrogel networks at elevated temperature, thereafter enhancing the permeability of microparticle scaffolds. Importantly, the hydrogel scaffolds are readily fabricated via on-chip quick gelation by triggered release of Ca2+ from the Ca-EDTA complex; it is also quite exciting that very mild release of microencapsulated cells is achieved via controlled degradation of hydrogel scaffolds through a simple strategy of competitive affinity of Ca2+ from the Ca-Alginate complex. This finding suggests that we are able to control cellular encapsulation and release through ion-induced gelation and degradation of the hydrogel scaffolds. Subsequently, we demonstrate a high viability of microencapsulated cells in the microgel scaffolds.

Highly flexible, tailorable and all-solid-state supercapacitors from carbon nanotube–MnOx composite films

Carbon nanotube (CNT) films are promising materials for constructing highly flexible composites for supercapacitor electrodes. Here, we prepared composite films with a sandwich structure and outstanding electrochemical properties by electrodepositing manganese oxide (MnOx) on the flexible CNT macrofilms. All-solid-state supercapacitors were then fabricated from the CNT–MnOx composite films using poly(vinylalcohol)–potassium hydroxide as the gel electrolyte. The supercapacitors have a high performance with a specific capacitance of 73.4 F g−1 and an energy density of 6.2 W h kg−1. The supercapacitors also exhibit high flexibility and stability under bending and kneading. When cut into small parts, the supercapacitor fragments can still work independently. The highly flexible and tailorable supercapacitors have great potential in flexible electronics in the future.

Self-Assemblies of Single-Walled Carbon Nanotubes through Tunable Tethering of Pyrenes by Dextrin for Rapidly Chiral Sensing

Pyrene-modified dextrin (Py-Dex) was synthesized via the Schiff base reaction between reducing end of dextrins and 1-aminopyrene, and then self-assemblies of single-walled carbon nanotubes (SWNTs) were fabricated through the tunable tethering of pyrene to SWNTs by dextrin chains. The Py-Dex-SWNTs assemblies were found to be significantly water-soluble because of the synergistic effect of dextrin chains and pyrene moieties. Py-Dex and Py-Dex-SWNTs were adequately characterized by NMR, UV-vis, fluorescence spectroscopy, Raman spectroscopy, matrix-assisted laser desorption/ionization-time of flight mass spectroscopy, and transmission electron microscopy. The tethering effect of dextrin toward pyrene moieties was clearly revealed and was found to be tunable by adjusting the length of dextrin chains. The fluorescence of pyrene moieties was sufficiently quenched by SWNTs with the support of dextrin chains. Furthermore, the Py-Dex-SWNTs assemblies were used for chiral selective sensing by introducing cyclodextrins as chiral binding sites. The rapid chiral sensing was successfully tested for different enantiomers.

Biocompatible Amphiphilic Hydrogel–Solid Dimer Particles as Colloidal Surfactants

Emulsions of two immiscible liquids can slowly coalesce over time when stabilized by surfactant molecules. Pickering emulsions stabilized by colloidal particles can be much more stable. Here, we fabricate biocompatible amphiphilic dimer particles using a hydrogel, a strongly hydrophilic material, and achieve large contrast in the wetting properties of the two bulbs, resulting in enhanced stabilization of emulsions. We generate monodisperse single emulsions of alginate and shellac solution in oil using a flow-focusing microfluidics device. Shellac precipitates from water and forms a solid bulb at the periphery of the droplet when the emulsion is exposed to acid. Molecular interactions result in amphiphilic dimer particles that consist of two joined bulbs: one hydrogel bulb of alginate in water and the other hydrophobic bulb of shellac. Alginate in the hydrogel compartment can be cross-linked using calcium cations to obtain stable particles. Analogous to surfactant molecules at the interface, the resultant amphiphilic particles stand at the water/oil interface with the hydrogel bulb submerged in water and the hydrophobic bulb in oil and are thus able to stabilize both water-in-oil and oil-in-water emulsions, making these amphiphilic hydrogel-solid particles ideal colloidal surfactants for various applications.