Microfluidic Devices and Methods for Use Thereof in Multicellular Assays of Secretion

Abstract

Methods and devices are provided herein for identifying a cell population comprising an effector cell that exerts an extracellular effect. In one embodiment the method comprises retaining in a microreactor a cell population comprising one or more effector cells, wherein the contents of the microreactor further comprise a readout particle population comprising one or more readout particles, incubating the cell population and the readout particle population within the microreactor, assaying the cell population for the presence of the extracellular effect, wherein the readout particle population or subpopulation thereof provides a direct or indirect readout of the extracellular effect, and determining, based on the results of the assaying step, whether one or more effector cells within the cell population exerts the extracellular effect on the readout particle. If an extracellular effect is measured, the cell population is recovered for further analysis to determine the cell or cells responsible for the effect.

1 - 218 . (canceled) 
     
     
         219 . A method of identifying an antibody secreting cell (ASC) that secretes a virus neutralizing antibody, comprising:
 retaining in a plurality of microreactors a plurality of cell populations, wherein individual cell populations of the plurality comprise one or more ASCs, wherein the contents of individual microreactors of the plurality of microreactors each comprise a readout particle population comprising one or more readout particles, and wherein the individual cell populations are retained in the individual microreactors;   introducing a plurality of accessory particle populations into the individual microreactors, wherein individual accessory particle populations of the plurality comprise a plurality of virus particles and the individual accessory particle populations are retained in individual microreactors;   incubating the individual cell populations, the readout particle populations and the accessory particle populations within the individual microreactors to provide secreted antibodies;   assaying the individual microreactors for the presence of a virus neutralizing antibody,   determining, based on the results of the assaying step, whether one or more of the cell populations comprises an ASC that secretes a virus neutralizing antibody.   
     
     
         220 . The method of  claim 219 , wherein the readout particle population comprises a homogeneous population of readout particles. 
     
     
         221 . The method of  claim 219 , wherein the readout particle population comprises a heterogeneous population of readout particles. 
     
     
         222 . The method of  claim 220 , wherein the homogeneous population of readout particles comprises a homogeneous population of readout cells. 
     
     
         223 . The method of  claim 221 , wherein the heterogeneous population of readout particles comprises a heterogeneous population of readout cells. 
     
     
         224 . The method of  claim 219 , wherein the readout particle population or subpopulation thereof is immobilized on a surface of the individual microreactors. 
     
     
         225 . The method of  claim 219 , further comprising maintaining the individual cell populations in substantially a single plane. 
     
     
         226 . The method of  claim 219 , further comprising maintaining the readout particle populations in substantially a single plane. 
     
     
         227 . The method of  claim 219 , wherein the plurality of accessory particles further comprise a fluorescent substrate, fluorophore, a secondary antibody, or a combination thereof. 
     
     
         228 . The method of  claim 219 , wherein the individual accessory particle populations of the plurality are heterogeneous accessory particle populations. 
     
     
         229 . The method of  claim 219 , wherein the plurality of virus particles is engineered to include a fluorescent protein expressed by a readout cell following virus infection of the readout cell. 
     
     
         230 . The method of  claim 219 , wherein assaying the individual microreactors comprises assaying the morphology of the readout particle populations. 
     
     
         231 . The method of  claim 219 , wherein assaying the individual microreactors comprises assaying binding of the secreted antibodies to the virus particles. 
     
     
         232 . The method of  claim 219 , wherein assaying the individual microreactors comprises assaying the expression of fluorescent proteins within the readout particle populations that are upregulated during viral infection. 
     
     
         233 . The method of  claim 219 , wherein assaying the individual microreactors comprises assaying the death of the readout particle populations. 
     
     
         234 . The method of  claim 219 , further comprising substantially isolating the individual microreactors from their surrounding environments. 
     
     
         235 . The method of  claim 219 , wherein if a cell population comprises an ASC that secretes a virus neutralizing antibody, the method further comprises recovering the cell population comprising the ASC that secretes the virus neutralizing antibody or a portion thereof to obtain a recovered cell population. 
     
     
         236 . The method of  claim 230 , wherein the recovering step comprises positioning the open end of a microcapillary in a microreactor comprising the cell population comprising the ASC that secretes the virus neutralizing antibody and aspirating the microreactor's contents or a portion thereof to obtain a recovered aspirated cell population. 
     
     
         237 . The method of  claim 236 , wherein the microcapillary is mounted on a robotic micromanipulation system on a microscope or the microcapillary is controlled robotically. 
     
     
         238 . The method of  claim 235 , further comprising,
 retaining a plurality of cell subpopulations originating from the recovered cell population in a plurality of vessels, wherein each cell subpopulation is present in an individual vessel,   lysing the individual cell subpopulations to provide lysed cell subpopulations, and   amplifying one or more nucleic acids within each of the lysed cell populations.