Methods and apparatus for the manipulation of particle suspensions and testing thereof

Abstract

Apparatus and methods are provided for analysis of individual particles in a microfluidic device. The methods involve the immobilization of an array of particles in suspension and the application of experimental compounds. Such methods can also include electrophysiology studies including patch clamp recording, electroporation, or both in the same microfluidic device. The apparatus provided includes a microfluidic device coupled to a multi-well structure and an interface for controlling the flow of media within the microchannel device.

What is claimed is: 
     
         1 . A method for performing an assay, the method comprising:
 loading a fluid comprising a suspension of particles into one or more input wells of a microfluidic device, the microfluidic device further comprising an output well and a channel fluidly connecting at least one input well of the one or more input wells and the output well;   coupling a pressure interface to the microfluidic device to create an air tight seal around the input well, the output well, or both;   applying pneumatic pressure to a gas-fluid interface in the input well to flow at least a portion of a fluid in the input well into the channel of the microfluidic device; and   imaging at least a portion of the channel of the microfluidic device after said applying pneumatic pressure to measure an adhesive interaction of the particles.   
     
     
         2 . The method of  claim 1 , wherein the adhesive interaction comprises adherence of at least some of the particles to the channel of the microfluidic device, adherence of at least some of the particles to a layer of particles attached to the channel, detachment of at least some of the particles from the channel, or combinations thereof. 
     
     
         3 . The method of  claim 1 , further comprising analyzing, using a controller, an image of the at least a portion of the channel of the microfluidic device to assess characteristics of particles within the channel. 
     
     
         4 . The method of  claim 3 , wherein said analyzing comprises measuring migration of the particles during the flow, morphology of the particles, and internalization of particles with a coating or within a gel provided in the microfluidic device. 
     
     
         5 . The method of  claim 3 , wherein the particles comprise cells, said analyzing comprising measuring cell growth, cell viability, genetic profiles, or expression profiles the cells or combinations thereof. 
     
     
         6 . The method of  claim 1 , wherein said applying pneumatic pressure includes applying pneumatic pressure sufficient to achieve a predetermined shear flow profile in the channel. 
     
     
         7 . The method of  claim 1 , further comprising stopping the flow prior to said imaging. 
     
     
         8 . The method of  claim 7 , wherein said stopping the flow comprises controlling the pneumatic pressure to cause the fluid to flow through the channel at a speed below 5 nm/ms. 
     
     
         9 . The method of  claim 1 , wherein said coupling the pressure interface includes applying mechanical pressure to deform a deformable layer of the pressure interface and create the air tight seal. 
     
     
         10 . The method of  claim 1 , wherein said coupling the pressure interface includes applying a negative pressure to one or more cavities of the microfluidic device fluidly decoupled from the input well and the output well to deform a deformable layer of the pressure interface and create the air tight seal. 
     
     
         11 . The method of  claim 1 , wherein flowing the fluid through the channel comprises flowing the fluid through a first channel and a second channel fluidly coupled to the input well, the method further comprising controlling a shear force applied to the fluid in the first channel and the second channel. 
     
     
         12 . The method of  claim 11 , wherein said controlling the shear force comprises:
 flowing a first portion of the fluid through a first portion of the first channel having a first cross-sectional area and flowing a second portion of the fluid through a first portion of the second channel having a second cross-sectional area different from the first cross-sectional area; and   flowing the first portion of the fluid through a second portion of the first channel having a third cross-sectional area and flowing the second portion of the fluid through a second portion of the second channel having a fourth cross-sectional area which is the same as the third cross-sectional area.   
     
     
         13 . The method of  claim 1  further comprising loading the fluid into the output well prior to said coupling the pressure interface to the microfluidic device. 
     
     
         14 . The method  claim 1  further comprising coating the channel with a monolayer of cells and loading the fluid into the input well after said coating the channel. 
     
     
         15 . The method of  claim 1  further comprising applying pneumatic pressure to a second gas-fluid interface in a second input well of the microfluidic device to flow at least another portion of a fluid in the second input well into the channel, the channel fluidly connecting the first and second input wells to the output well. 
     
     
         16 . The method of  claim 1  further comprising reversing a direction of flow through the channel. 
     
     
         17 . The method of  claim 1  further comprising:
 generating, using a computer system, control signals for applying pneumatic pressure corresponding to a user-specified flow or shear rate profile. 
 
     
     
         18 . The method of  claim 17  further comprising:
 receiving the user-specified flow or shear rate profile in the computer system coupled to the pressure interface, wherein the user-specified flow or shear rate profile comprises a profile as a function of time. 
 
     
     
         19 . A method for performing a biofilm assay, the method comprising:
 loading a bacterial suspension into an input well of a microfluidic device, the microfluidic device further comprising an output well and a channel fluidly connecting the input and output wells;   coupling a pressure interface to the microfluidic device to create an air tight seal around the input well, the output well, or both;   applying pneumatic pressure to a gas-fluid interface in the input well to flow at least a portion of the bacterial suspension in the input well into the channel such that a bacterial biofilm forms on a surface of the channel, and wherein applying the pneumatic pressure includes applying a shear profile to the bacterial biofilm;   imaging at least a portion of the channel of the microfluidic device after biofilm formation; and   measuring changes in biofilm properties due to either shear force or compound interaction.   
     
     
         20 . A method for performing an assay, the method comprising:
 treating a channel of a microfluidic device with a biomolecule or establishing a cell monolayer on at least one surface of the channel, the channel fluidly connecting an input well and an output well of the microfluidic device;   loading a suspension of blood cells into the input wells of the microfluidic device   coupling a pressure interface to the microfluidic device to create an air tight seal around the input well, the output well, or both;   applying pneumatic pressure to a gas-fluid interface in the input well to flow at least a portion of the suspension in the input well into the channel;   applying a shear profile resulting in movement of the blood cells across the at least one surface of the channel;   imaging at least a portion of the channel of the microfluidic device; and   measuring an adhesion of the blood cells in response to either a shear force profile or presence of a compound in the channel.   
     
     
         21 . The method of  claim 20 , wherein the suspension of blood cells comprises whole blood, treated whole blood, or a whole blood fraction, and wherein the adhesion of the blood cells is platelet adhesion or a thrombus formation inside the channel. 
     
     
         22 . The method of  claim 20 , wherein the blood cells comprise leukocytes, neutrophil, or both, and wherein said measuring an adhesion of blood cells comprises measuring static or rolling adhesion of the leukocytes or neutrophils.