Single cell capture with polymer capture films

Abstract

The present invention provides methods, systems, assemblies, and articles for capturing single cells with a polymer capture film. In certain embodiments, the polymer capture films comprise a plurality of individual channels with top and bottom openings, where the channels are dimensioned such that a single cell is: i) is captured inside the channel, partially or substantially occluding the channel, when negative pressure is provided to the bottom opening; or ii) is captured by the top opening, but does not enter the channel, when negative pressure is provided to the bottom opening. In some embodiments, the channels of the polymer capture film align with the wells of a multi-well chip such that the cell, or the contents of the single cell, may be transferred to a corresponding well.

We claim: 
     
         1 . An article of manufacture comprising: a polymer capture film, wherein said polymer capture film comprises a top surface, a bottom surface, and a plurality of individual channels extending through said polymer capture film, wherein each of said individual channels:
 a) has a top opening in said top surface of said polymer capture film,   b) has a bottom opening in said bottom surface of said polymer capture film, and   c) is dimensioned such that one cell, and only said one cell, from a cell suspension:
 i) is captured inside said individual channel, partially or substantially occluding said individual channel, when negative pressure is provided to said bottom opening; or 
 ii) is captured by said top opening, but does not enter said individual channel, when negative pressure is provided to said bottom opening. 
   
     
     
         2 . The article of manufacture of  claim 1 , wherein said polymer capture film comprises an inert and/or a hydrophobic polymer. 
     
     
         3 . The article of manufacture of  claim 1 , further comprising at least one film guide component attached to, or integral with, said polymer capture film, wherein said film guide component is configured to allow alignment of said plurality of individual channels in said polymer capture film one-for-one with wells in a multi-well device and/or holes in a polymer film support plate. 
     
     
         4 . The article of manufacture of  claim 1 , wherein said top openings of said plurality of individual channels matches one-for-one, and aligns with, the well openings in a multi-well device. 
     
     
         5 . The article of manufacture of  claim 1 , wherein said plurality of individual channels comprises at least 500 individual channels. 
     
     
         6 . The article of manufacture of  claim 1 , wherein each of said individual channels is dimensioned such that one cell, and only said one cell, from a cell suspension is captured by said top opening, but does not enter said individual channel, when negative pressure is provided to said bottom opening. 
     
     
         7 . The article of manufacture of  claim 1 , wherein said polymer film has a thickness between 10 μm and 50 μm, and wherein said top surface has an area between 1 cm 2  and 30 cm 2 . 
     
     
         8 . The article of manufacture of  claim 1 , wherein each of said individual channels has a diameter of about 2-6 μm. 
     
     
         9 . A system comprising:
 a) a polymer capture film comprising a top surface, a bottom surface, and a plurality of individual channels extending through said polymer capture film, wherein each of said individual channels:
 i) has a top opening in said top surface of said polymer capture film, 
 ii) has a bottom opening in said bottom surface of said polymer capture film, and 
 iii) is dimensioned such that one cell, and only said one cell, from a cell suspension:
 A) is captured inside said individual channel, partially or substantially occluding said individual channel, when negative pressure is provided to said bottom opening; or 
 B) is captured by said top opening, but does not enter said individual channel, when negative pressure is provided to said bottom opening; and 
 
   b) a first component selected from the group consisting of:
 i) a porous layer comprising a porous material that allows liquid, but not cells, to pass therethrough, wherein said porous layer is dimensioned to be contacted with said bottom surface of said polymer capture film such that said bottom opening of each of said plurality of individuals channels is covered by said porous material; 
 ii) a film support component comprising a substrate with a plurality of individual holes therethrough which align one-for-one with said plurality of individual channels in said polymer capture film, wherein said film support component is dimensioned to be contacted with, and support, said bottom surface of said polymer capture film; 
 iii) a multi-well chip comprising a plurality of individual wells, 
   wherein said multi-well chip is dimensioned to be contacted with said top surface of said polymer capture film such that said plurality of individual wells align one-for-one with said plurality of individual channels in said polymer capture film; and
 iv) a cell that: A) is captured inside one of said individual channels, or B) is captured by said top opening of one of said individual channels. 
   
     
     
         10 . The system of  claim 9 , further comprising a housing base, wherein said housing base is configured to contain said polymer capture film and comprises at least one base feature selected from the group consisting of: a vacuum connection port, at least two guide pins for aligning said polymer capture film and said film support component, a film support plate recess, a liquid reservoir, a gasket, and at least one connection rod for connection to a housing top. 
     
     
         11 . The system of  claim 9 , further comprising a housing top, wherein said housing top is configured to connect to a housing base to enclose said polymer capture film and comprises at least one top feature selected from the group consisting of: an inlet port, an outlet port, a slot array, a guide pin receiver, and a connection rod receiver. 
     
     
         12 . The system of  claim 9 , wherein said polymer capture film comprises an inert and/or hydrophobic polymer. 
     
     
         13 . The system of Claim , wherein said polymer capture film further comprises at least one film guide component attached to, or integral with, said polymer capture film, wherein said film guide component is configured to allow alignment of said plurality of individual channels in said polymer capture film one-for-one with said wells in said multi-well device and/or said holes in said polymer film support plate. 
     
     
         14 . The system of  claim 9 , wherein said top openings of said plurality of individual channels matches one-for-one, and aligns with, said well openings in said multi-well device and/or said holes in said polymer film support plate. 
     
     
         15 . The system of  claim 9 , wherein said polymer capture film has at least one of the properties selected from the group consisting of: i) said plurality of individual channels comprises at least 500 individual channels, ii) each of said individual channels is dimensioned such that one cell, and only said one cell, from a cell suspension is captured by said top opening, but does not enter said individual channel, when negative pressure is provided to said bottom opening, iii) has a thickness between 10 μm and 50 μm, iv) said top surface has an area between 1 cm 2  and 30 cm 2 ; and iv) each of said individual channels has a diameter of about 2-6 μm. 
     
     
         16 . The system of  claim 9 , wherein said cell is selected from the group consisting of: a platelet, a red blood cell, a neutrophil, a lymphocyte, an exocrine cell, a fibroblast, an osteocyte, a chondrocyte or a liver cell, a goblet or ciliated cell, a macrophage, a hematopoietic stem cell, an adipocyte filled with stored lipid, and a neuron. 
     
     
         17 . The system of  claim 1 , further comprising a second component selected from the group consisting of: a vacuum pump that provides said negative pressure, a centrifuge motor configured to spin a housing base, a diaphragm pump, a chip alignment component, a swing arm attached to a swing arm support rod, a sample delivery component, a user interface display, and at least one waste container. 
     
     
         18 . A method of isolating single cells comprising:
 a) contacting a cell suspension with a cell-isolating system, wherein said cell-isolating system comprises:
 i) a polymer capture film comprising a top surface, a bottom surface, and a plurality of individual channels extending through said polymer capture film, wherein each of said individual channels:
 A) has a top opening in said top surface of said polymer capture film, 
 B) has a bottom opening in said bottom surface of said polymer capture film, and 
 C) is dimensioned such that one cell, and only said one cell, from said cell suspension:
 1) is captured inside said individual channel, partially or substantially occluding said individual channel, when negative pressure is provided to said bottom opening; or 
 2) is captured by said top opening, but does not enter said individual channel, when negative pressure is provided to said bottom opening; and 
 
 
 ii) a pneumatic device that provides said negative pressure, and 
   b) incubating said cell suspension with said cell-isolating system, wherein said pneumatic device provides said negative pressure, such that at least some of said individual channels in said polymer captures film captures one cell from said cell suspension such that at least some of said plurality of individual channels has one captured cell; and   c) washing said polymer capture film to remove non-captured cells to generate a captured-cell polymer film.   
     
     
         19 . The method of  claim 18 , further comprising bringing together said captured-cell polymer film and a multi-well device to generate an assembly, wherein said multi-well device has a plurality of well-openings, and wherein said top openings of said plurality of individual channels of said captured-cells polymer film matches one-for-one, and aligns with, said plurality of well openings in said multi-well device. 
     
     
         20 . The method of  claim 18 , wherein said cell isolating system further comprises: i)a porous layer comprising a porous material that allows liquid, but not cells, to pass therethrough, wherein said porous layer is dimensioned to be contacted with said bottom surface of said polymer capture film such that said bottom opening of each of said plurality of individuals channels is covered by said porous material; or ii) a film support component comprising a substrate with a plurality of individual holes therethrough which align one-for-one with said plurality of individual channels in said polymer capture film, wherein said film support component is dimensioned to be contacted with, and support, said bottom surface of said polymer capture film.