SYSTEM, FLUIDICS CARTRIDGE, AND METHODS FOR USING ACTUATED SURFACE-ATTACHED POSTS FOR PROCESSING CELLS

1 . A cell processing system comprising:

a fluidics cartridge comprising:

a cell processing chamber comprising a bottom substrate and a top substrate separated by the gap, wherein the cell processing chamber further comprises a micropost array, wherein the micropost array comprises a plurality of surface-attached microposts arranged on a micropost substrate, and wherein the micropost substrate is positioned atop the bottom substrate; and

a control instrument;

wherein the surface-attached posts are configured for actuation in the presence of an actuation force, wherein no binding agents are disposed on or integrated with the surface-attached posts, the bottom substrate, the top substrate, or the micropost substrate, and wherein the bottom substrate and the top substrate are arranged atop a registration feature configured for mounting on the control instrument.

2 . The cell processing system of claim 1 , wherein the fluidics cartridge further comprises one or more sample reservoirs, one or more wash reservoirs, one or more supply cell processing chambers, one or more waste reservoirs, and one or more eluent reservoirs fluidly connected via an arrangement of fluid channels to the cell processing chamber.

3 . The cell processing system of claim 2 , wherein a fluid control port is provided in each of the fluid channels.

4 . The cell processing system of claim 3 , wherein the one or more sample reservoirs, the one or more wash reservoirs, the one or more supply cell processing chambers, the one or more waste reservoirs, and the one or more eluent reservoirs each comprise an inlet and an outlet.

5 . The cell processing system of claim 4 , wherein a fluid control port is provided at the outlet of the sample reservoir, a fluid control port is provided at the outlet of the wash reservoir, a fluid control port is provided at the inlet of the waste reservoir, and a fluid control port is provided at the inlet of the eluent reservoir.

6 . The cell processing system of claim 5 , wherein the fluid control ports comprise pinch valves.

7 . The cell processing system of claim 6 , wherein a first pump is fluidly connected to the sample reservoir and a second pump is fluidly connected to the wash reservoir.

8 . The cell processing system of claim 7 , wherein the first pump and the second pump are capable of supplying positive pressure and negative pressure to the cell processing chamber.

9 . The cell processing system of any one of claims 2 to 8 , wherein one or more of the sample reservoir, the wash reservoir, the waste reservoir, and the eluent reservoir comprise seals that are gas permeable but not liquid permeable.

10 . The cell processing system of any one of claims 2 to 9 , wherein the control instrument comprises a base that houses one or more mechanisms for providing one or more actuation forces to the microposts, one or more mechanisms for counting cells in the cell processing chamber, one or more pneumatics for pumping and controlling fluids in the fluidics cartridge, and a controller.

11 . The cell processing system of claim 10 , wherein the actuation force is selected from the group consisting of a magnetic field, a thermal field, a sonic field, an optical field, an electrical field, and a vibrational field.

12 . The cell processing system of any one of claims 1 to 11 , wherein the control instrument comprises a platform configured to interface with the fluidics cartridge.

13 . The cell processing system of claim 12 , wherein the platform comprises a plurality of fluid control ports positioned to correspond to the fluid channels of the fluidics cartridge.

14 . The cell processing system of claim 13 , wherein each of the fluid control ports comprise a valve mechanism.

15 . The cell processing system of claim 14 , wherein the valve mechanism is a pinch valve.

16 . The cell processing system of any one of claims 12 to 15 , wherein the platform further comprises an optical window substantially aligned with the cell processing chamber of the fluidics cartridge.

17 . The cell processing system of claim 16 , wherein the one or more mechanisms for counting cells in the cell processing chamber is an optical imaging system.

18 . The cell processing system of any one of claims 1 to 17 , wherein the cell processing system is a standalone device.

19 . The cell processing system of any one of claims 1 to 17 , wherein the cell processing system further comprises an automated robotics system for processing biological materials.

20 . The cell processing system of claim 19 , wherein the automated robotics system for processing biological materials comprises a multi-well plate.

21 . The cell processing system of claim 20 , wherein the multi-well plate is selected from the group consisting of a 12-well plate, a 24-well plate, and a 96-well plate.

22 . The cell processing system of claim 20 or claim 21 , wherein dimensions of the registration feature of the fluidics cartridge substantially correspond to dimensions of the multi-well plate.

23 . The cell processing system of any one of claims 20 to 22 , wherein the automated robotics system for processing biological materials further comprises one or more pipettes for processing fluids from the multi-well plate.

24 . The cell processing system of any one of claims 20 to 23 , wherein the automated robotics system for processing biological materials further comprises a pipette for processing fluids from the eluent reservoir of the fluidics cartridge.

25 . The cell processing system of any one of claims 10 to 24 , wherein the one or more mechanisms for counting cells in the cell processing chamber comprises measurement of electrical resistance, flow cytometry, image analysis, spectrophotometry, detection of fluorescence of fluorescently labeled cells, or combinations thereof.

26 . The cell processing system of any one of claims 1 to 25 , wherein the microposts are formed of polydimethylsiloxane (PDMS).

27 . The cell processing system of any one of claims 1 to 26 , wherein the microposts range in length from about 1 μm to about 100 μm.

28 . The cell processing system of any one of claims 1 to 27 , wherein the microposts range in diameter from about 0.1 μm to about 10 μm.

29 . The cell processing system of any one of claims 1 to 28 , wherein the microposts have a cross-sectional shape selected from the group consisting of circular, ovular, square, rectangular, and triangular.

30 . The cell processing system of any one of claims 1 to 29 , wherein the microposts are oriented substantially normal to the plane of the substrate.

31 . The cell processing system of any one of claims 1 to 29 , wherein the microposts are oriented at an angle α with respect to normal of the plane of the substrate.

32 . The cell processing system of any one of claims 1 to 29 , wherein the microposts are oriented at a pitch of from about 0 μm to about 50 μm.

33 . The cell processing system of any one of claims 1 to 32 , further comprising a controller capable of executing program instructions.

34 . The cell processing system of any one of claims 1 to 33 , further comprising a user interface.

35 . The cell processing system of any one of claims 1 to 34 , further comprising a communications interface.

36 . The cell processing system of any one of claims 1 to 35 , further comprising a power source.

37 . A method for processing cells comprising the use of a cell processing system of any one of claims 1 to 36 , comprising the steps of:

(a) introducing a sample fluid to the sample reservoir, wherein the sample fluid comprises cells, and wherein the cells are suspended in the sample fluid;

(b) flowing the sample fluid into the cell processing chamber;

(c) precipitating the cells suspended in the sample fluid onto the micropost substrate amongst the surface-attached microposts, wherein no actuation forces are applied to the surface-attached microposts;

(d) performing a cell wash cycle comprising flowing wash buffer solution out of the wash reservoir, through cell processing chamber, and into the waste reservoir, wherein the cells remain precipitated onto the micropost substrate amongst the surface-attached microposts, and wherein no actuation forces are applied to the surface-attached microposts;

(e) repeating step (d) as needed to wash the cells precipitated onto the micropost substrate amongst the surface-attached microposts;

(f) performing a cell recovery cycle comprising flowing wash buffer solution through the cell processing chamber, wherein actuation forces are applied to the surface-attached microposts to resuspend the cells into the flowing wash buffer solution, thereby producing a cell-containing eluent; and

(g) flowing the cell-containing eluent into the eluent reservoir.

38 . A method for processing cells comprising the use of a cell processing system of any one of claims 1 to 36 , comprising the steps of:

(a) introducing a sample fluid to the sample reservoir, wherein the sample fluid comprises cells, and wherein the cells are suspended in the sample fluid;

(b) flowing the sample fluid into the cell processing chamber;

(c) precipitating the cells suspended in the sample fluid onto the micropost substrate amongst the surface-attached microposts, wherein no actuation forces are applied to the surface-attached microposts;

(d) performing a cell wash cycle comprising flowing wash buffer solution out of the wash reservoir, through cell processing chamber, and into the waste reservoir, wherein the cells remain precipitated onto the micropost substrate amongst the surface-attached microposts, and wherein no actuation forces are applied to the surface-attached microposts;

(e) repeating step (d) as needed to wash the cells precipitated onto the micropost substrate amongst the surface-attached microposts;

(f) performing a cell counting operation with the cell counting mechanism to determine the number of cells in the cell processing chamber;

(g) performing a cell recovery cycle comprising flowing wash buffer solution through the cell processing chamber, wherein actuation forces are applied to the surface-attached microposts to resuspend the cells into the flowing wash buffer solution, thereby producing a cell-containing eluent; and

(h) flowing the cell-containing eluent into the eluent reservoir.

39 . A method for processing cells comprising the use of a cell processing system of any one of claims 1 to 36 , comprising the steps of:

(a) introducing a sample fluid to the sample reservoir, wherein the sample fluid comprises cells, and wherein the cells are suspended in the sample fluid;

(b) flowing the sample fluid into the cell processing chamber;

(c) precipitating the cells suspended in the sample fluid onto the micropost substrate amongst the surface-attached microposts, wherein no actuation forces are applied to the surface-attached microposts;

(d) performing a cell lysis cycle, wherein actuation forces are applied to the surface-attached microposts to produce a beating motion by the surface-attached microposts, thereby producing a lysed cell-containing eluent; and

(e) flowing the lysed cell-containing eluent into the eluent reservoir.

40 . A method for processing cells comprising the use of a cell processing system of any one of claims 1 to 36 , comprising the steps of:

(a) introducing a sample fluid to the sample reservoir, wherein the sample fluid comprises cells, and wherein the cells are suspended in the sample fluid;

(b) flowing the sample fluid into the cell processing chamber while the surface-attached microposts are actuated, wherein the sample fluid is flowed at rate slow enough that the cells are not pushed out of the cell processing chamber;

(c) performing a cell wash cycle comprising flowing wash buffer solution out of the wash reservoir into the cell processing chamber, wherein actuation forces are applied to the surface-attached microposts, and further wherein the wash buffer solution is flowed at a rate slow enough that the cells are not pushed out of the cell processing chamber;

(d) performing a cell culture cycle comprising flowing cell culture media out of the cell culture media reservoir into the cell processing chamber, wherein actuation forces are applied to the surface-attached microposts, and further wherein the cell culture media is flowed at a rate slow enough that the cells are not pushed out of the cell processing chamber;

(e) performing a cell recovery cycle comprising flowing wash buffer solution through the cell processing chamber, wherein actuation forces are applied to the surface-attached microposts, and further wherein the wash buffer solution is flowed at a rate fast enough that the cells are not pushed out of the cell processing chamber; and

(f) flowing the cell-containing eluent into the eluent reservoir.

41 . A method for processing cells comprising the use of a cell processing system of any one of claims 1 to 36 , comprising the steps of:

(a) introducing a sample fluid to the sample reservoir, wherein the sample fluid comprises cells, and wherein the cells are suspended in the sample fluid;

(b) flowing the sample fluid into the cell processing chamber;

(c) precipitating the cells suspended in the sample fluid onto the micropost substrate amongst the surface-attached microposts, wherein no actuation forces are applied to the surface-attached microposts;

(d) performing a cell wash cycle comprising flowing wash buffer solution out of the wash reservoir, through cell processing chamber, and into the waste reservoir, wherein the cells remain precipitated onto the micropost substrate amongst the surface-attached microposts, and wherein no actuation forces are applied to the surface-attached microposts;

(e) repeating step (d) as needed to wash the cells precipitated onto the micropost substrate amongst the surface-attached microposts;

(f) performing a cell culture cycle by flowing cell culture media into cell processing chamber while microposts are not actuated and providing time and conditions necessary for cell growth, expansion, and maintenance;

(g) performing a cell recovery cycle comprising flowing wash buffer solution through the cell processing chamber, wherein actuation forces are applied to the surface-attached microposts to resuspend the cells into the flowing wash buffer solution, thereby producing a cell-containing eluent; and

(h) flowing the cell-containing eluent into the eluent reservoir.

42 . The method of claim 41 or 42 , further comprising the step of performing a cell counting operation with the cell counting mechanism to determine the number of cells in the cell processing chamber, wherein the cell counting operation is performed before the cell recovery cycle step.

43 . The method of any one of claims 37 to 42 , wherein the sample fluid comprises cells comprising clumps of cells, wherein the cells comprising clumps of cells are suspended in the sample fluid, and wherein step (b) further comprises applying actuation forces to the surface-attached microposts to break up the clumps of cells.

44 . The method of any one of claims 37 to 43 , wherein prior to step (a), the sample fluid is produced by a cell concentration process comprising centrifuging a sample comprising cells to produce a cell pellet, followed by resuspending cells in the cell pellet in solution to produce the sample fluid.

45 . The cell processing system of any one of claims 1 to 35 , further comprising a microarray on the top substrate opposing the microposts, wherein the microarray is functionalized with analyte capture elements.

46 . A method for processing cells comprising the use of a cell processing system of claim 45 , comprising the steps of:

(a) introducing a sample fluid to the sample reservoir, wherein the sample fluid comprises cells, and wherein the cells are suspended in the sample fluid;

(b) flowing the sample fluid into the cell processing chamber;

(c) precipitating the cells suspended in the sample fluid onto the micropost substrate amongst the surface-attached microposts, wherein no actuation forces are applied to the surface-attached microposts;

(d) flowing a lysis buffer into the cell processing chamber, thereby producing lysed cells and analytes; and

(e) applying actuation forces to the surface-attached microposts to mix the lysed cells and analytes in the cell processing chamber, wherein analytes bind to the analyte capture elements of the microarray.

47 . The cell processing system of any one of claims 1 to 35 , further comprising a cell concentration module.

48 . A method for processing cells comprising the use of a cell processing system of claim 47 , comprising the steps of:

(a) introducing a sample fluid to the sample reservoir, wherein the sample fluid comprises cells, and wherein the cells are suspended in the sample fluid;

(b) flowing the sample fluid into the cell concentration module and performing a cell concentration process to produce a concentrated sample fluid; and

(c) flowing the concentrated sample fluid into the cell processing chamber for further processing.

49 . The cell processing system of any one of claims 1 to 35 , wherein the micropost array comprises a flow path formed by the absence of microposts.

50 . The cell processing system of claim 49 , wherein at least some portion of the flow path is curved and configured to aggregate cells at the outside of curves such that the cells precipitate and/or are pushed into the microposts.

51 . The cell processing system of claim 50 , wherein the flow path is serpentine-shaped.

52 . The cell processing system of claim 50 , wherein the flow path is spiral-shaped.

53 . A method of making a cell processing system of any one of claims 47 to 52 , wherein the method comprises fabricating the micropost array in a high density and using a tool to crush unwanted microposts to form the flow path.

54 . The cell processing system of any one of claims 1 to 35 , wherein the micropost array comprises arrangements of micropost barriers configured to trap cells such that the cells precipitate and/or are pushed into the microposts.

55 . The cell processing system of claim 54 , wherein the micropost barriers are arc-shaped, U-shaped, V-shaped, or bar-shaped.

56 . The cell processing system of any one of claims 1 to 35 , further comprising features on the top substrate opposing the microposts, wherein the features are configured to assist cells to precipitate out of solution and/or facilitate microfluidic cell separation.

57 . The cell processing system of claim 55 , wherein the features are arranged in a herringbone configuration.

58 . The cell processing system of any one of claims 1 to 35 , further comprising one or more electrodes provided in the bottom substrate, the top substrate, or both the bottom substrate and the top substrate.