Microchip Based On Biochemical Concentration Gradients And Temperature Gradients And Its Application

Exploring the influence of cellular microenvironment on cell physiological activities is a hot topic in the field of cell biology.Cells in living organisms are regulated by various physical and biochemical factors.The physical factors include the temperature,density,stiffness of ECM(extracellular matrix),the shear force and pressure on cells,etc.The biochemical factors include growth factor that regulates the cell behavior.An integrated micro-device which provides biochemical concentration gradients and temperature gradients simultaneously was developed in this paper.Cells remained vigorous when planted in the microchip and were influenced by dual-factor gradients.The construction of the platform enables the dual gradients loading on cells in the field of microfluidics,and is of great significance for exploring the response of various cells to two gradients.Firstly,the PDMS microchip containing concentration gradients generation(CGG)module and cell culture array was designed.The tree network was adopted to generate 5 concentrations.Through COMSOL simulation analysis,the qualitative experiment of red and blue ink and the quantitative experiment of Rhodamine 123 fluorescent dye,taking into account the chip fabrication,the size of 45 ?m deep 150 ?m wide and a perfusion rate of 20 ?L/h were adopted in subsequent studies.Cell culture array was connected to CGG module though a 5 ?m-high microstructure.Then the ITO microchip generating temperature gradients was designed.According to the simulation of temperature distribution on the ITO glass surface,the shape of the resistive film and the glass substrate were optimized,and a slender,hollow-shaped ITO temperature gradient chip was designed to generate uniform temperature gradients on ITO glass.The temperature on the ITO microchip was measured by infrared thermal imaging and fluorescence imaging.A gradient of 37.6/38.6/39.3/40.2/40.3? was obtained.An experiment to verify the validation of the integrated platform was carried out.PC12 cells were planted in PDMS microchip.An NGF concentration gradient of 0/25/50/75/100 ng/mL and a temperature gradient of 37.6/38.6/39.3/40.2/40.3? were loaded on the attached cells simultaneously.The statistics of PC12 neurite length illustrated that PC12 cells were influenced by the two factors and differentiated in different level.At a result,the experiment verify the validation of the integrated platform.In summary,this study realized the parallel loading of multiple temperatures on the microfluidic platform and the simultaneous loading of concentration gradient and temperature gradient,which provided an effective experimental platform for studying the effects of two factors on physiological activities of cells.