Methods and systems for microfluidic screening

Abstract

Provided are methods and systems useful for screening large libraries of effector molecules. Such methods and systems are particularly useful in microfluidic systems and devices. The methods and systems provided herein utilize encoded effectors to screen large libraries of effectors.

1 .- 20 . (canceled) 
     
     
         21 . A screening system comprising:
 (a) a microfluidic device comprising a droplet formation region connected to a flow path, wherein the droplet formation region is configured to generate a plurality of droplets, and wherein the flow path is configured to incubate the plurality of droplets for at least 14 minutes with a dispersion ratio of less than 4%;   (b) a plurality of unique scaffolds encapsulated in the plurality of droplets wherein a subset of the plurality of droplets each comprises at least one unique scaffold of the plurality of unique scaffolds, and wherein at least a subset of the plurality of the unique scaffolds each comprises:
 (i) an effector bound to the unique scaffold via a cleavable linker and configured to be released from the scaffold upon cleavage of the cleavable linker to generate a released effector in a droplet of the subset of the plurality of droplets; and, 
 (ii) a barcode corresponding to and identifying the effector; and 
   (c) at least one of: a sorting device for sorting a droplet of the plurality of droplets or an injection device for injecting a reagent into a droplet of the plurality of droplets, based on the signal.   
     
     
         22 . The system of  claim 21 , wherein the droplet formation region is configured to generate droplets at a frequency of at least 70 Hz. 
     
     
         23 . The system of  claim 21 , wherein the plurality of unique scaffolds comprise at least 80,000 unique scaffolds, wherein each unique scaffold comprises a unique effector. 
     
     
         24 . The system of  claim 21 , wherein the effector comprises at least one effector subunit, the barcode comprises at least one barcode subunit, and the barcode subunit corresponds to and identifies the effector or the effector subunit. 
     
     
         25 . The system of  claim 21 , wherein the barcode comprises a plurality of barcode subunits, wherein the effector comprises a plurality of effector subunits, wherein one or more barcode subunits of the plurality of barcode subunits corresponds to and identify one effector subunit of the plurality of effector subunits. 
     
     
         26 . The system of  claim 21 , wherein the barcode is a nucleic acid barcode. 
     
     
         27 . The system of  claim 26 , wherein the nucleic acid barcode comprises a sequence unique to the effector. 
     
     
         28 . The system of  claim 26 , wherein the nucleic acid barcode comprises a sequence unique to the unique scaffold. 
     
     
         29 . The system of  claim 21 , wherein the barcode is deoxyribonucleic acid (DNA), ribonucleic acid (RNA), a peptide, or a peptide nucleic acid. 
     
     
         30 . The system of  claim 21 , wherein the effector is selected from the group consisting of: a compound, a protein, a peptide, a nucleic acid molecule, an enzyme, a small molecule, a small molecule fragment, and a drug. 
     
     
         31 . The system of claim  1 , wherein the scaffold is at least 1 micrometer (μM) in diameter. 
     
     
         32 . The system of claim  1 , wherein the scaffold comprises a bead selected from the group consisting of a polymer bead, a glass bead, a metal bead, and a magnetic bead. 
     
     
         33 . The system of claim  1 , wherein the unique scaffold comprises a bead comprising a polymer resin. 
     
     
         34 . The system of  claim 33 , wherein the polymer resin comprises polyethylene glycol. 
     
     
         35 . The system of  claim 33 , wherein the polymer resin comprises reactive groups for attachment of functionalities. 
     
     
         36 . The system of  claim 33 , wherein the bead comprises an internal section and an outer surface, wherein the internal section comprises a plurality of reactive groups and the bead surface comprises a surface modification rendering the bead surface orthogonally protected to the reactive groups. 
     
     
         37 . The system of  claim 21 , wherein the concentration of the released effector in each droplet of the subset of plurality of droplets is within a predetermined range of values. 
     
     
         38 . The system of  claim 21 , wherein the concentrations of the released effectors in the subset of the plurality of droplets differ no more than 2-fold. 
     
     
         39 . The system of  claim 21 , wherein the barcode is a nucleic acid barcode covalently bound to the scaffold. 
     
     
         40 . The system of  claim 21 , wherein the barcode comprises information related to the synthesis history of the effector.