Methods and Devices for Analyzing Particles

Abstract

Methods, devices and systems for analyzing precious samples of cells, including single cells are provided. The methods, devices, and systems in various embodiments of the invention are used to assess genomic heterogeneity, which has been recognized as a central feature of many cancers and plays a critical role in disease initiation, progression, and response to treatment. The methods devices and systems are also used to analyze embryonic biopsies for preimplantation genetic diagnosis (PGD). In one embodiment, the devices, systems and methods provided herein allow for the construction of genomic and RNA-seq libraries without a pre-amplification step.

1 . A variable volume chamber, the chamber comprising:
 (a) a flow layer defining a reaction chamber and flow channels, wherein at least one wall of the reaction chamber or a portion thereof is an elastomeric membrane, the membrane having:
 (i) a neutral position; and 
 (ii) a plurality of expanded positions; 
   wherein the expanded positions determine the volume of the reaction chamber, whereby the volume of the reaction chamber is V at the neutral position and the volume of the reaction chamber is greater than V in the plurality of expanded positions, wherein V is less than 1 μl, and wherein the volume at a maximally expanded position is equal to or greater than 2×V; and   (b) a blank layer to seal the flow layer.   
     
     
         2 . The variable volume chamber of  claim 1 , wherein the chamber further comprises a control layer adjacent the flow layer or separated from the control layer by a membrane layer, wherein the control layer defines control lines. 
     
     
         3 . The variable volume chamber of  claim 1 , wherein the elastomeric membrane also has a plurality of reduced positions whereby the volume is less than V in the plurality of reduced positions, and wherein the reduced volume is equal to or less than 0.5×V. 
     
     
         4 . The variable volume chamber of  claim 1 , wherein the chamber further comprises a pressure chamber that surrounds the variable volume chamber or chambers, whereby the pressure chamber is operable to produce a variable pressure on the exterior surface of the elastomeric membrane. 
     
     
         5 . The variable volume chamber of  claim 1 , wherein the chamber further comprises a displacement chamber, whereby the displacement chamber and the reaction chamber are aligned, and whereby the reaction chamber is operable to expand into the displacement chamber and the displacement chamber is operable to produce a positive pressure on the exterior surface of the elastomeric membrane of the reaction chamber. 
     
     
         6 . The variable volume chamber of  claim 1 , wherein the volume at the expanded position is equal to or greater than: (a) 3×V; (b) 4×V; (c) 5×V; (d) 6×V; (e) 7×V; (f) 8×V; (g) 9×V; (h) 10×V; (i) 11×V; (j) 12×V; (k) 13×V; (1) 14×V; (m) 15×V; (n) 16×V; (o) 17×V; (p); 18×V; (q) 19×V; (r) 20×V; (s) 25×V; (t) 30×V; (u) 35×V; (v) 40×V; (w) 45×V; (x) 50×V; (y) 55×V; (z) 60×V; (aa) 65×V; (bb); 70×V; (cc) 75×V; (dd) 80×V; (ee) 85×V; (ff) 90×V; (gg) 95×V; (hh) or is equal to 100×V. 
     
     
         7 . The variable volume chamber of  claim 3 , wherein the volume at the reduced position is equal to or less than: (a) 0.4×V; (b) 0.3×V; (c) 0.2×V; (d) 0.1×V; (e) 0.09×V; (f) 0.08×V; (g) 0.07×V; (h) 0.06×V; (i) 0.05×V; (j) 0.04×V; (k) 0.03×V; (1) 0.02×V; (m) 0.01×V; or is essentially zero×V. 
     
     
         8 . The variable volume chamber of  claim 1 , wherein the expanded position is equal to or greater than 5×V and the reduced position is equal to or less than 0.5×V. 
     
     
         9 . The variable volume chamber of  claim 1 , wherein the expanded position is equal to or greater than 3×V and the reduced position is equal to or less than 0.4×V. 
     
     
         10 . The variable volume chamber of  claim 1 , wherein the expanded position is equal to or greater than 10×V and the reduced position is equal to or less than 0.5×V. 
     
     
         11 . A variable volume chamber, the chamber comprising:
 (a) a flow layer defining a reaction chamber and flow channels, wherein at least one wall of the reaction chamber or a portion thereof is an elastomeric membrane, the membrane having:
 (i) a neutral position; and 
 (ii) a plurality of expanded positions; 
   wherein the expanded positions determine the volume of the reaction chamber, whereby the volume of the reaction chamber is V at the neutral position and the volume of the reaction chamber is greater than V in the plurality of expanded positions, wherein V is less than 1 μl, and wherein the volume at a maximally expanded position is equal to or greater than 2×V; and   (b) a blank layer to seal the flow layer; and   (c) a displacement chamber that surrounds the reaction chamber, whereby the displacement chamber and the reaction chamber are aligned, such that the reaction chamber is operable to expand into the displacement chamber and wherein the displacement chamber is operable to constrain the expansion of the reaction chamber.   
     
     
         12 . The variable volume chamber of  claim 11 , wherein the chamber further comprises a control layer adjacent the flow layer or separated from the control layer by a membrane layer, wherein the control layer defines control lines. 
     
     
         13 . The variable volume chamber of  claim 11 , wherein the elastomeric membrane also has a plurality of reduced positions whereby the volume is less than V in the plurality of reduced positions, and wherein the reduced volume is equal to or less than 0.5×V. 
     
     
         14 . The variable volume chamber of  claim 11 , wherein the chamber further comprises a pressure chamber that surrounds the variable volume chamber or chambers, whereby the pressure chamber is operable to produce a variable pressure on the exterior surface of the elastomeric membrane. 
     
     
         15 . The variable volume chamber of  claim 11 , wherein the volume at the expanded position is equal to or greater than: (a) 3×V; (b) 4×V; (c) 5×V; (d) 6×V; (e) 7×V; (f) 8×V; (g) 9×V; (h) 10×V; (i) 11×V; (j) 12×V; (k) 13×V; (1) 14×V; (m) 15×V; (n) 16×V; (o) 17×V; (p); 18×V; (q) 19×V; (r) 20×V; (s) 25×V; (t) 30×V; (u) 35×V; (v) 40×V; (w) 45×V; (x) 50×V; (y) 55×V; (z) 60×V; (aa) 65×V; (bb); 70×V; (cc) 75×V; (dd) 80×V; (ee) 85×V; (ff) 90×V; (gg) 95×V; (hh) or is equal to 100×V. 
     
     
         16 . The variable volume chamber of  claim 13 , wherein the volume at the reduced position is equal to or less than: (a) 0.4×V; (b) 0.3×V; (c) 0.2×V; (d) 0.1×V; (e) 0.09×V; (f) 0.08×V; (g) 0.07×V; (h) 0.06×V; (i) 0.05×V; (j) 0.04×V; (k) 0.03×V; (1) 0.02×V; (m) 0.01×V; or is essentially zero×V. 
     
     
         17 . The variable volume chamber of  claim 11 , wherein the expanded position is equal to or greater than 5×V and the reduced position is equal to or less than 0.5×V. 
     
     
         18 . The variable volume chamber of  claim 11 , wherein the expanded position is equal to or greater than 3×V and the reduced position is equal to or less than 0.4×V. 
     
     
         19 . The variable volume chamber of  claim 11 , wherein the expanded position is equal to or greater than 10×V and the reduced position is equal to or less than 0.5×V. 
     
     
         20 . A variable volume chamber, the chamber comprising:
 (a) a flow layer defining a reaction chamber and flow channels, wherein at least one wall of the reaction chamber or a portion thereof is an elastomeric membrane, the membrane having:
 (i) a neutral position; 
 (ii) a plurality of expanded positions; and 
 (iii) a plurality of reduced positions; 
   wherein the expanded and reduced positions determine the volume of the reaction chamber, whereby the volume of the reaction chamber is V at the neutral position, the volume of the reaction chamber is greater than V in the plurality of expanded positions and the volume is less than V in the plurality of reduced positions, such that the volume at a maximally reduced position is essentially zero and wherein V is less than 1 μl;   (b) a control layer adjacent the flow layer defining control lines;   (c) a seal layer adjacent the control, to seal the flow; and   (d) a pressure chamber that surrounds the variable volume chamber or chambers, whereby the pressure chamber is operable to produce a positive pressure on the exterior surface of the elastomeric membrane and wherein the displacement chamber is operable to constrain the expansion of the reaction chamber.   
     
     
         21 . The variable volume chamber of  claim 1 , wherein the flow layer is a polymeric organosilicon compound. 
     
     
         22 . The variable volume chamber of  claim 1 , wherein the flow layer is polydimethylsiloxane (PDMS). 
     
     
         23 . The variable volume chamber of  claim 1 , wherein the chamber further comprises oligonucleotides deposited in the reaction chamber. 
     
     
         24 . The variable volume chamber of  claim 23 , wherein the oligonucleotides are indexing oligonucleotides. 
     
     
         25 . The variable volume chamber of  claim 1 , wherein the reaction chamber is in fluid communication with an auxiliary chamber. 
     
     
         26 . The variable volume chamber of  claim 25 , wherein the auxiliary chamber further comprises deposited oligonucleotides which may be released into the reaction chamber. 
     
     
         27 . The variable volume chamber of  claim 1 , wherein the reaction chamber further comprises a cell trap. 
     
     
         28 . The variable volume chamber of  claim 1 , wherein the chamber further comprises microparticles deposited in the reaction chamber. 
     
     
         29 . The variable volume chamber of  claim 28 , wherein the microparticles are capture beads. 
     
     
         30 .- 121 . (canceled)