The Application Of Quantum Dots Encoded Microspheres In Microfluidics Flow Cytometry System

In chapter 1, some new progresses in development of micro-fluidic-chipanalysis systems at home and abroad are introduced systematically. The progressesin the aspects of miniaturization of instrument, application to the biologicalanalysis, micromachining technology for chips and various detection techniquesdeveloped are viewed especially. And some problems faced now are presented.Flow cytometry is a general method for analyzing microparticles such as cells,bacteria and other microorganism with high efficiency, integrating optics, fluidics,electronics, and computer technology. With the application of all kinds ofadvanced technologies, flow cytometers are getting smaller, cheaper, faster, moreintegrated and easier in performance. In this chapter, Quantum dots (QD) is reviewed, its basic knowledge andcharacters,synthesis,labeling approach,recent progress and prospect in potentialapplication to life science,especially to immunoassay and rapid diagnosticanalysis. Quantum dots (QD),with a diameter of 1-100 nanometers in size,as aspecific kind of nanoparticles developed in recent years,provide an extremelygood fluorescent signal and then can well be used for fluorescent labeling,also asan ideal bioconjugated fluorescent probe for bioanalysis,because of their uniqueoptical and electronic properties,such as broad excitation spectrum,narrowemission spectrum,good tune and neglectable optical bleach phenomenon etc.Although its research and application has just been explored,QD have attractedgreat attention from scientists in many fields.In chapter 2, first, a novel method of synthesizing QDs fluorescence-encodedmicrospheres by dispersing polymerization technique is reported. A series ofpolystyrene microspheres from 1 μm to 5 μm in diameter are attained. By thefollowing one-step swelling procedure, the different nanocrystals were carried intothe inner of Ps microspheres quantificationally. As-prepared fluorescence-encodedmicrospheres have the following outstanding advantages: the nanocrystals are notabsorbed on the surface of the microspheres but carried into the inner ofmicrospheres, so the NCs cannot be isolated or leaked out easily. Besides,compared with microspheres loaded by traditional organic dyes, as-synthesizedphotoluminescent microspheres have better photostability, and it is much moredifficult for them to be photobleached. Moreover, this method can be operatedeasily and reproducibly. Second, the method of tagging cells with quantum dots byelectroporation is developed. An electroporation method is proposed to make QDspass through the membranes of K562 leukaemia cells. This method is operatedsimply and facilitates the tagging experiment greatly. This method brings forwardsome more extrusive advantages than cell endocytosis and microinjection method,and there should be a very good foreground for further applications.In chapter 2, the microchip flow cytometer and the method of detectingbiochemical sample are studied. Applying electrokinetic focusing technology, thewhole fluidic controlling system is set up on a microchip with four electrodes in it.By using laser induced fluorescence system, the prototype instrument is small, andthe operation is simple. Then, a microfluidic chip is designed and fabricated, andin coupled with flow cytometry. Using Na2PO4 solution added with hydroxypropylmethyl cellulose (HPMC) as buffer and sheathing fluid, it solves a series ofproblems met during the flowing of fluorescent microspheres in the microchannel,so as to keep the fluidic conditions under more effective control. By using laserinduced fluorescence system assembled in the lab and applying electrokineticfocusing technology, the counting of the microspheres is achieved. And real timeobservation of their actual fluidic state through an inverted fluorescencemicroscope is made. The whole system is compact and easy to be operated. Theusage of reagent and sample is little and the speed of the assay is quick.Combining microspheres and flow cytometry, the method of detecting epidemichemorrhagic fever and new-castle disease is developed. It also set the foundationfor the research of detecting multiple animal diseases simultaneously.