Smartphone Based Microfluidic Lab-on-Chip Device For Real-Time Detection,Counting And Sizing Of Living Algae

Global economic integration and the growth of transnational trade volume promote the growth of international shipping scale.However,the discharge of untreated ballast water from ships can cause serious biological invasion and pose a great threat to the local marine ecological environment.The International Ballast Water Convention specifies the concentrations of active organisms with different sizes in ballast water which meets discharge requirements.Microalgae is the main component of phytoplankton in ballast water,so it is very important to analyze the activity and concentration of microalgae rapidly and accurately.The existing ballast water detection methods are mainly indicative analysis method on site and detailed analysis method in laboratory.Laboratory analysis is time-consuming and bulky,and on-site analysis is only indicative analysis whose detection accuracy is not high enough.Therefore,it is necessary to develop a portable device for detailed analysis of activity of microalgae in ballast water.The main research contents are:(1)Taking the theory of chlorophyll fluorescence detection as the basis for judging the activity of microalgae,microfluidic chip as the sample detection platform,and smartphone as the data analysis platform,designs a smartphone-based fluorescent imaging system for on-site and real-time detection,counting and sizing of living algae in ballast water samples.The device consists of a sample-carrying platform,a tiny peristaltic pump,a microfluidic chip,a customer-made fluorescence microscopic imaging platform and a smartphone.When the living algae is driven through the excitation light lighting area of the microfluidic chip by the micro peristaltic pump,the chlorophyll in the living algae will stimulate chlorophyll fluorescence,which is captured by the smartphone camera after filtration and amplification and finally displayed on the phone screen in the form of red fluorescent dot.(2)Taking the computer vision technology as the basis for counting the microalgae,develops an algorithm that can detect,count and calculate the size of living microalgae,and transplants the algorithm to Android phone.The algorithm is divided into three parts:detection,tracking and counting.Based on Inter-Frame Difference Method,the detection algorithm performs subtraction,binarization,morphology closed operation,contour detection and convex hull calculation of two consecutive frames in the detection video.Each convex hull represents a detected target.The tracking algorithm is based on centroid tracking.The prediction part of the tracking algorithm predicts the target’s tracking box in the current frame by computing the weighted sum of the differences between the adjacent abscissas(or ordinates)of n consecutive positions in the trajectory.The match part of the tracking algorithm is based on Euclidean distance.The counting algorithm sets up a counting line in the difference frame.When the tracked target crosses the counting line,the counting value is added by one.(3)Designs several detection experiments,including the living chlorella detection experiment,the mixed sample of living chlorella and tetraselmis detection experiment,the mixed sample of dead and living Chlorella detection experiment and the size calibration verification experiment,to verify the identification ability and counting accuracy of the device for living microalgae of different sizes.The results show that for the detection device,the detection accuracy of living chlorella is 92%,the accuracy of sizing is 94%,and the device is able to judge the activity of microalgae.(4)Designs an imaging system which can show light field and fluorescence field at the same time on the basis of the above system.The beam containing both blue light and fluorescence is split by a spectroscope,so that the fluorescence is collected by the Android phone camera in the horizontal direction,and the blue light is collected by the USB camera in the vertical direction.The USB camera is connected to another Android phone through an adapter.Two android phones display both blue and fluorescence field images of microalgae samples at the same time.In this thesis,chlorophyll fluorescence is used as the theory of microalgae activity judgment,microfluidic chip is used as the sample detection platform,smart phone is used as data analysis platform,a microscopic analysis method for activity of microalgae in ballast water is designed,which provides a new idea for detailed on-site analysis of microalgae in ballast water.