Study On The Detection Of Plant Immune Elicitors Based On Droplet-based Microfluidics

Droplet microfluidic chip is a science and technology of the micro scale space for fluid manipulation as the main characteristics,it has a very wide range of uses in biology,chemistry and other aspects.Nitric oxide(NO),Calcium ion(Ca2+),Hydrogen peroxide(H2O2)are an important signaling molecule in plant.It plays an important role in a variety plant physiological process,such as growth,development and environmental stimuli.This paper mainly uses the platform of droplet microfluidic chip to achieve the detection of fluorescence in droplets,the fluorescence was produced by BY-2 cells with fluorescent probe which has been induced by Chitosan Oligosaccharide(COS).Droplet microfluidic provides a reference for the study of carbohydrate plant immune elicitors by use of microfluidics.The main contents were as follows:We designed and fabricated a droplet microfluidic chip which can encapsulate the single BY-2 cell,the droplet size was determined by adjusting the water phase flow and oil phase flow.The single BY-2 cell in droplets were counted and we found that the probability of single cell fit Poisson distribution.In this experiment,BY-2 cells in the exponential growth phases were loaded fluorescence probes,fluorescence probes including DAF-FM DA,Fluo-3AM and H2DCF-DA.BY-2 cells with the fluorescence probe were incubated for a period time.were then stimulated by 50?g/mL COS solution,the significant fluorescence was observed by fluorescence microscope.BY-2 cells with fluorescence were arranged into suitable concentration and then be encapsulated by the droplet microfluidic chip,the cells in droplets can also produce significant fluorescence.The BY-2 cell with fluorescence probe were induced by COS,and then were incubated in 96 well plates(black)and droplets,we used fluorescent labelling system to detect the fluorescence of BY-2 cells in 96 well plates(black)and droplets,then found that the fluorescence intensity variation trend is consistent.It proved that the droplet can be used as a carrying device to detect the fluorescence signal which induced by carbohydrate plant immune elicitors.This is the first study to combined plant cells with droplet microfluidic technology,it provides a new platform to analysis single plant cell,and provides a reference for the high-throughput screening of carbohydrate plant immune elicitors by use of microfluidics.