NEXT GENERATION OPTICALLY ACTIVATED LARGE PARTICLE SORTING

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Several avenues of research have a critical need for high speed sorting of large particles, including the selection of whole organisms, multicellular particles and the selection of molecules from combinatorial libraries synthesized on large particles. High speed sorting has to date primarily relied on charge based droplet sorting coupled with multiparameter optical analysis of particles in a flow cytometer, which is an indispensable technique for numerous biomedical applications including rare cell isolation, chromosome sorting and cellular display molecular selection among many others. However, droplet based flow sorters have significant limitations when large particles are considered. First, increasing particle size requires largerorifices to prevent clogging and effects on droplet break off points. Sorting orifices suffer increasing turbulence as their diameter increases, which requires the use of lower linear velocities and restricts sorting rates (<1000/s). This, effectively limits particle size (<100 ¿m) and has led to alternative large particle sorting approaches, including mechanical stream diversion and micro-channel fluidic switching. Our effort focuseson solutions to large particle sorting that will result in the development of sorters that will sort particles up to 1 mm in diameter at rates comparable to current conventional droplet based sorters (>104/s). To accomplish this, we will target each technical limitation of current sorting technology with unique solutions for large particle systems. First, we will develop high speed synchronous particle delivery systems to overcome the statistical uncertainty of particle delivery and maximize sorting rates by providing known particle positions for sorting events. Second, we will leverage our recent low cost flow cytometry developments in lasers and data acquisition systems to create inexpensive low linear velocity parallel flow cytometry analyzers to maximize particle throughput. Third, we will create droplet on demand sorters that are not limited by particle size. While these technical developments will be most applicable to large particle sorting, they will also have ancillary benefits to conventional sorting, as they will dramatically reduce system cost and create valuable parallel analysis technology for high speed sorting. Finally, we will construct high-speedlarge particle sorters for internal testing and key external collaborations to sort large particles at high rates for selection of aptamers and peptides as well as rare tumor microspheroid collection.