•The effect of ion concentration on vesicle formation was investigated on a designed miniaturized reactor.•Na+ and Cl− would bind with the lipid head and act a much strong hydrophobic repulsion on the lipid tail.•High salt concentration enhanced hydrophobic repulsion on lipids increasingly, and forced lipids attached firmly on the substrate.•Much larger external disturbance would be needed for vesicle formation in salt solutions than that in pure water.Lipid vesicle formation is known to be suppressed in salt solutions, but the mechanism of this phenomenon remains unclear. In order to better understand this issue, the effect of salt concentrations (0–800 mM) of sodium chloride on the behavior of L-α-phosphatidylcholine (PC) in aqueous solution was investigated in this work. The results showed that fusion among vesicles, micelles and bilayers may be essential for vesicle formation. With addition of ions and an increase in ion concentration, the lipids became constrained in lateral movement and packed increasingly tightly. The resulted hard supported phospholipid bilayers (SPBs) were thus more difficult to detach from the substrate to form vesicles. These phenomena were tried to be explained at molecular level. Hydrophobic effect is the original cause of lipid vesicle formation, which in fact is absence of attraction between the involved substances. That is to say, the stronger the 3D network was bounded in the medium, the stronger the hydrophobic repulsion on the lipids would be. This might be one reason for the suppression of vesicle formation in salt solution.Download high-res image (108KB)Download full-size image

•Controllable electroporation of cells was achieved using microcavity electrodes.•An optimized microcavity electrode for cell electroporation/electro-fusion was obtained.•A cell electroporation model was developed and validated.•High-yield electro-fusion in a microfluidic device with microcavity electrodes was achieved.Cell electro-fusion includes four steps, cell alignment, cell electroporation, reconstruction of cytomembrane and cytoplasm exchange. The cell alignment and electroporation steps are highly related to the intensity and distribution of the electric field, which depend on the applied voltage as well as the microelectrode structure. The microelectrode structures were first evaluated based on the numerical analysis of the electric field and the transmembrane potential induced on biological cells when the cell electroporation and electro-fusion were performed based on different designs of microelectrodes. Microelectrodes in a micro-cavity geometry can induce electroporation around the contact area of the paired cells for high-yield electro-fusion. Microfluidic chips with co-planar microelectrodes and microelectrodes within micro-cavities have been fabricated and tested for electro-fusion of Myoblast cells, and the experimental results confirmed the numerical analysis.Download high-res image (128KB)Download full-size imageControllable cell electroporation is successfully achieved by microcavity electrodes.

A microfluidic chip with preparation module and collection module for giant vesicles by microfluidic technology and microelectrode array was fabricated. First, lipid solution was loaded into the microelectrode array through microchannels to form lipid film, then an electric field was subsequently loaded on the microelectrode array, and giant vesicles with controlled diameter were formed efficiently. The ratio of formed stable giant spherical vesicles could reach up to 60%. Giant vesicles and other materials were flushed into the upper layer of the collection chamber by microchannel. 90% of the stable giant spherical vesicles with 10–50 mm diameter could be sorted in the upper layer of the collection chamber by using micropore filter and gravity depositing. This microfluidic chip could overcome some defects existing in the current preparation method such as low efficiency, wide distribution of diameters, as well as difficult for screening and collection.In this study, a microfluidic chip with the preparation module and collection module was designed and fabricated. The microfluidic technique and microelectrode array were used for the preparation and collection of giant vesicles. The stable giant spherical vesicles were efficiently formed, sorted and collected in this chip.

•Surface morphology of lipid films is accurately and rapidly characterized.•A SPR imaging device based on an angle interrogation manner is constructed.•Mathematical model is developed to describe the shift of the light path.Surface topographies of lipid films have an important significance in the analysis of the preparation of giant unilamellar vesicles (GUVs). In order to achieve accurately high-throughput and rapidly analysis of surface topographies of lipid films, a homemade SPR imaging device is constructed based on the classical Kretschmann configuration and an angle interrogation manner. A mathematical model is developed to accurately describe the shift including the light path in different conditions and the change of the illumination point on the CCD camera, and thus a SPR curve for each sampling point can also be achieved, based on this calculation method. The experiment results show that the topographies of lipid films formed in distinct experimental conditions can be accurately characterized, and the measuring resolution of the thickness lipid film may reach 0.05 nm. Compared with existing SPRi devices, which realize detection by monitoring the change of the reflective-light intensity, this new SPRi system can achieve the change of the resonance angle on the entire sensing surface. Thus, it has higher detection accuracy as the traditional angle-interrogation SPR sensor, with much wider detectable range of refractive index.

•Microstructures can be fabricated within enclosed microchannels.•2D patterns and 3D microstructures can be formed.•Formed microstructures and patterns can be modified easily.•Magnetic-beads-based microstructure can be used for cell biosensors.•This method is much convenient for chip design and fabrication.A large number of microscale structures have been used to elaborate flowing control or complex biological and chemical reaction on microfluidic chips. However, it is still inconvenient to fabricate microstructures with different heights (or depths) on the same substrate. These kinds of microstructures can be fabricated by using the photolithography and wet-etching method step by step, but involves time-consuming design and fabrication process, as well as complicated alignment of different masters. In addition, few existing methods can be used to perform fabrication within enclosed microfluidic networks. It is also difficult to change or remove existing microstructures within these networks. In this study, a magnetic-beads-based approach is presented to build microstructures in enclosed microfluidic networks. Electromagnetic field generated by microfabricated conducting wires (coils) is used to manipulate and trap magnetic beads on the bottom surface of a microchannel. These trapped beads are accumulated to form a microscale pile with desired shape, which can adjust liquid flow, dock cells, modify surface, and do some other things as those fabricated microstructures. Once the electromagnetic field is changed, trapped beads may form new shapes or be removed by a liquid flow. Besides being used in microfabrication, this magnetic-beads-based method can be used for novel microfluidic manipulation. It has been validated by forming microscale dam structure for cell docking and modified surface for cell patterning, as well as guiding the growth of neurons.

To study the impact of different temperatures on the growth and proliferation of myoblasts, specific electric fields were loaded on indium tin oxide (ITO) glass chips to generate temperature profiles. In several zones with different temperatures (38, 39, 40 and 41 °C), microchambers with equal size were fabricated for the culture of mouse myoblasts. Impacts of temperatures on growth and proliferation of myoblasts were studied by stimulating cells for continuous 5 days (30 min/day). Microscopic observation of cell morphology and flow cytometric detection results showed that certain thermal stimulation could promote the proliferation of myoblasts. The number of cells was obviously increased after stimulation at 40 °C. For equal total stimulation time, short-term and multiple thermal stimulation could achieve better effect, and maximum proliferation index could reach 38.39.Specific electric fields were loaded on ITO glass chips to generate temperature profiles to study their impact on the growth and proliferation of myoblasts. Cells were stimulated for continuous 5 days, and the results showed that certain thermal stimulation could promote the proliferation of myoblasts. Short-term and multiple thermal stimulation could achieve better effect.

A new cell electrofusion microfluidic chip with 19,000 pairs of micro-cavity structures patterned on vertical sidewalls of a serpentine-shaped microchannel has been designed and fabricated. In each micro-cavity structure, the two sidewalls perpendicular to the microchannel are made of SiO2 insulator, and that parallel to the microchannel is made of silicon as the microelectrode. One purpose of the design with micro-cavity microelectrode array is to obtain high membrane voltage occurring at the contact point of two paired cells, where cell fusion takes place. The device was tested to electrofuse NIH3T3 and myoblast cells under a relatively low voltage (~9 V). Under an AC electric field applied between the pair of microelectrodes positioned in the opposite micro-cavities, about 85–90 % micro-cavities captured cells, and about 60 % micro-cavities are effectively capable of trapping the desired two-cell pairs. DC electric pulses of low voltage (~9 V) were subsequently applied between the micro-cavity microelectrode arrays to induce electrofusion. Due to the concentration of the local electric field near the micro-cavity structure, fusion efficiency reaches about 50 % of total cells loaded into the device. Multi-cell electrofusion and membrane rupture at the end of cell chains are eliminated through the present novel design.

Large liposomes were prepared by using soybean lecithin and the co-evaporation method, and they were electrofused on a microelctrode-array based microchip. In the electrofusion process, the liposomes were aligned by the dielectrophoretic force, electroporated by using high-strength electric pulses, and subsequently fused by using continuous dielectrophoretic force. The experimental results showed that 20% liposomes could be fused on the chip, and the glass substrate and low depth-width ratio channel were helpful for the observation and control of the liposome fusion.

Cell-electrofusion chip technique is a rapidly developing cell-fusion method in the last decade. It has been widely used in the basic research and application in many fields such as genetics, distant cross breeding of animal and plant, developmental biology, immunology, medicine, food, and agriculture. The cell-electrofusion chip technique draws much attention due to its characteristics of high controllability, easy implementation, high efficiency, and harmless, as well as easy to be observed with low consumption. This article reviews the basic principle, method and development of this technique, and predicts its improving trends.

A microfluidic pool structure for cell docking and rapid mixing is described. The pool structure is defined as a microchamber on one structural layer of a bilayer chip and connects with two or more individual microchannels on the other structural layer. In contrast to the turbulent flow in a macroscale pool, laminar streams enter and exit this microfluidic pool structure with definite and controllable direction that may be influenced by the location and geometry of the pool. A simple microfluidic model was used to validate this hypothesis. In this model, a microscale pool structure was made on the lower layer of a chip and connected with three parallel microchannels in the upper layer. Simulation and experimental results indicated that the flow profile within the pool structure was determined by its geometry and location. This could be used as a flow control method and it was simpler than designs based on microvalve, hydraulic pressure, or electrokinetic force, and has some important applications. For example, controllable streams within this structure were used to immobilize biological cells along the microchannel walls. When different solution streams flowed through the pool, rapid diffusion of analytes occurred for short diffusion distance between vertical flow laminas. Furthermore, desired dilution (mixing) ratio could be obtained by controlling the geometry of the microfluidic pool.