System and methods

Abstract

We describe instruments, and cartridges therefor, for processing droplets of water in oil-in-water emulsions, the droplets containing biological entities such as cells. In one embodiment a method of such processing comprises: providing a plurality of the entities in a fluid: preparing a droplet from said fluid: determining whether said droplet contains one or more entities of said plurality of entities, or whether said droplet does not contain a said entity; sorting said droplet dependent on an outcome of said determination; and dispensing said sorted droplet into a reservoir, wherein said dispensing comprises: identifying said sorted droplet for dispensing; extracting said sorted droplet from a first fluidic flow path of said fluid by transferring said sorted droplet from said first fluidic flow path into a second fluidic flow path; and ejecting said sorted droplet from said second fluidic flow path into said reservoir by applying pressure to said second fluidic flow path. In preferred embodiments the droplet contents are tracked so that the contents of an individual droplet can be sorted, selectively dispensed, and afterwards retrieved for further processing or analysis, or for subsequent use, for example in fermenting or the like.

1 . A microfluidic cartridge for automated sorting and dispensing of sorted microfluidic droplets, the cartridge comprising:
 a sorting input channel coupled to a sorting region, said sorting region comprising two or more sorting output channels coupled to said input channel and a droplet director to selectively direct a droplet from said sorting input channel to a selected one of said sorting output channels; and   a droplet dispenser coupled to one of said sorting output channels to dispense selected said droplets from said cartridge for collection in a reservoir, wherein said droplet dispenser comprises a droplet ejection mechanism to eject droplets from an emulsion flow into a dispenser output channel for collection in said reservoir.   
     
     
         2 . The microfluidic cartridge as claimed in  claim 1  further comprising an emulsion flow buffer region between said sorting region and said dispensing region to control an emulsion flow rate to said droplet dispenser. 
     
     
         3 . The microfluidic cartridge as claimed in  claim 1 , further comprising a droplet incubation region having droplet input, a droplet holding region to hold droplets for incubation, and an output coupled to said sorting input channel. 
     
     
         4 . The microfluidic cartridge as claimed in  claim 1 , wherein said dispenser output channel includes a mechanism to promote liberation of a droplet or the contents of a droplet from said emulsion. 
     
     
         5 . The microfluidic cartridge as claimed in  claim 1 , wherein said droplet dispenser is configured for selecting droplets with a defined number or numeric range of biological entities per droplet. 
     
     
         6 . The microfluidic cartridge as claimed in  claim 1 , further comprising a plurality of sensors for detecting a said droplet at different locations of said microfluidic cartridge at different points in time to determine a velocity of said droplet in said microfluidic cartridge, and wherein said droplet director is configured to direct a said droplet dependent on an outcome of said velocity determination. 
     
     
         7 . The microfluidic cartridge as claimed in  claim 1 , wherein said droplet ejection mechanism comprises a mechanism to increase a pressure in said emulsion flow to direct droplets into said dispenser output channel, in particular wherein said droplet ejection mechanism is configured to selectively modulate said pressure to select a droplet in said emulsion flow for ejection. 
     
     
         8 . The microfluidic cartridge as claimed in  claim 1 , wherein one or more emulsion flow buffer regions are provided between the sorting region and the droplet dispenser, to control an emulsion flow rate to the droplet dispenser. 
     
     
         9 . A microdroplet processing system including the microfluidic cartridge as claimed in  claim 8  further comprising a sensor to sense one or both of a speed and a position of said selected droplet, and a controller to modulate said pressure responsive to said speed/position. 
     
     
         10 . A microdroplet processing system comprising the microfluidic cartridge as claimed in  claim 1 , the microdroplet processing system further comprising a plurality of said reservoirs and a control system to selectively direct a dispensed droplet to a reservoir. 
     
     
         11 . The microdroplet processing system as claimed in  claim 9 , further comprising a plurality of said reservoirs and a control system to selectively direct a dispensed droplet to a reservoir. 
     
     
         12 . The microdroplet processing system as claimed in  10  further comprising a droplet sensing system, in particular associated with said sorting region, to sense one or more properties of a droplet for sorting or sorted into a said output channel; and
 wherein said control system is configured to store droplet data relating to said one or more sensed properties such that the droplet data for an individual droplet is retrievable to retrieve data for an individual droplet in an identified reservoir. 
 
     
     
         13 . A microfluidic system for providing a droplet containing one or more entities including the microfluidic cartridge as claimed in  claim 1 , the system further comprising:
 a droplet formation device for preparing a droplet from a fluid which contains a plurality of entities;   an analyser for determining whether a said droplet contains one or more entities of said plurality of entities;   wherein said sorting region of said microfluidic cartridge is configured for sorting a said droplet dependent on an outcome of said determination; and   a decoupler for decoupling a said sorted droplet from a fluidic flow path of said fluid in said microfluidic system, wherein said droplet dispenser of said microfluidic cartridge is configured for dispensing the decoupled droplet.   
     
     
         14 . The microfluidic system as claimed in  claim 13  wherein said droplet dispenser comprises a reservoir for storing a growth media fluid, and wherein said droplet dispenser is configured to dispense a said decoupled droplet in said growth media fluid. 
     
     
         15 . The microfluidic system as claimed in  claim 9 , wherein said sorting region or said droplet dispenser is configured to rotate a turntable comprising portions of first and second microfluidic flows to selectively direct droplets to one of first and second output microfluidic flows. 
     
     
         16 . The microfluidic cartridge as claimed in  claim 1 , wherein said sorting region or said droplet dispenser is configured to rotate a turntable comprising portions of first and second microfluidic flows to selectively direct droplets to one of first and second output microfluidic flows. 
     
     
         17 . A microfluidic system for providing one or more target entities in a droplet, the microfluidic system comprising:
 a first detector for detecting one or both of:
 whether a said droplet contains a said target entity or target entities; and 
 a property of a said target entity or target entities; 
   and wherein:   said sorting region is configured for sorting a said droplet dependent on an outcome of said detection; and   said droplet dispenser is configured for dispensing a said sorted droplet at one or more target locations, wherein said microfluidic system is configured to correlate a said target location with a said property of a said target entity or target entities detected in said first detector.   
     
     
         18 . An instrument for microfluidic droplet-based processing of biological entities, the instrument comprising:
 a droplet generation system to generate one or more water-in-oil emulsions of droplets comprising biological entities;   a droplet processing system to process said droplets; and   a droplet dispensing system to dispense processed said droplets into one or more reservoirs;   wherein said droplet processing system is configurable for multiple different modes of operation including at least a monoclonality mode of operation and an assay mode of operation.   
     
     
         19 . The instrument as claimed in  claim 18 , wherein said droplet processing system includes at least a droplet sort unit, wherein said monoclonality mode of operation comprises sorting droplets based on occupancy of droplets by said biological entities, and wherein said assay mode of operation includes incubating said droplets and comprises sorting droplets based on a product of said biological entities during said incubation. 
     
     
         20 . The instrument as claimed in  claim 18 , wherein said droplet processing system comprises one or more interchangeable or modular droplet processing cartridges. 
     
     
         21 . The instrument as claimed in  claim 20 , wherein said one or more interchangeable or modular droplet processing cartridges further comprise at least one droplet fusion unit, and wherein said modes of operation include a droplet fusion assay mode of operation.