A Microfluidic Oscillator Based Long-Range Gradient Generator And Its Application In Biology

Dilution generation plays an important role in many biological applications,such as enzyme-linked immunosorbent assays,drug screening,and environmental toxicology.Nowadays,dilution process is mainly dependent on either manual operation or robotic automation.For manual dilution process,it requires a lot of laborious manual operations,which is easy to make mistakes.For robotic dilution process,researchers have to buy an expensive sample loading equipment,which increases the cost of experiments.Microfluidic chip is a technique that controls the fluid at the micron scale.It has several features,such as small volume,fast detection and automated operation.In this study,we develop a novel microfluidic mixer based on microfluidic technology and fluid oscillation and a valve controlled microfluidic gradient generator,which can automatically generate a long-range concentration gradient of samples.We believe this microfluidic system provides a new platform for the researches of drug screening and environmental toxicology.The main contents of this study are as follows:1.This paper presents a RC circuit like microfluidic oscillator.This oscillator is simple in design and can achieve a rapid mixing of two or more different micro-scale liquids.The effect of different experimental parameters such as the applied pressure,the size of the air chamber and the oscillation frequency on the microfluidic oscillator are studied by theoretical analysis and experimental verification.This study provides a reference for subsequent chip design.2.We develop several peripheral devices,including a customized equipment for on-chip valve control,a high-density microfluidic pin-header connector and some expansion accessories.This equipment can be used to control the micro-valves and oscillate fluids on the chip.3.We develop a microsphere immune chip based on the microfluidic oscillator and the peripheral devices.This chip combines the advantages of fluid oscillation and microsphere liquid suspension reaction.It can enhance the mixing of the antibody and the antigen to reduce the time required for immunizations.After the reaction,the microspheres are further aggregated by a dam microstructure to improve the sensitivity of the detection.We used rabbit antibodies and FICT labeled goat anti-rabbit antibodies as the experimental model.By using our system,the detect limitation was 375 ng/ml,and the time required for immunization was decreased to 5min.4.We present a microfluidic long-range dilution chip for drug screening.To verify the function of the chip,we developed two serial dilution chips with the dilution rates of 1:1 and 1:4.We demonstrated that these chips have good reproducibility and the maximum dilution concentration of them was 1:3125.If the dilution level on the chip was increased,the dilution range could be further increased.In order to verify the biological application of the chip,the kinetic parameters of?-galactosidase were determined by this chip.The results showed that the K_m=603±73?M and k_cat=72±12/sec.In addition,this study used this chip to carry out the acute oxidative stress test of C.elegans,which provided a new platform for the study of the model bio-nematode.In this study,a microfluidic oscillator is proposed,and a long-range gradient generator is constructed based on the on-chip micro-valve technology and the external control device.We demonstrate that this microfluidic oscillator allows rapid mixing of liquids at microscale and promotes the reaction of enzymes and substrates as well as antigenic antibodies.We also demonstrate that a gradient generator that integrates this microfluidic oscillator can form a long-range of sample concentration gradients and can be used for the research of enzymatic reaction kinetics and C.elegans.