# US 9,435,765 B2 · Cartridge and system for manipulating samples in liquid droplets > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 9,435,765 B2 - Document ID: US9435765B2 - Status: Granted - Kind: B2 - Application number: 13/737,183 - Filed: 2013-01-09 - Published: 2013-05-16 - Granted: 2016-09-06 - Earliest claimed: 2011-07-22 - Assignees: Tecan Trading AG - Applicants: Tecan Trading AG - Inventors: S. Nicholas Reimitz; Bruce Richardson; Marc N. Feiglin; Anne R. Kopf-Sill - Web page: https://patentgrep.com/patent/US9435765B2 ## CPC - G01N 27/44782: {of a plurality of samples} - B01L 3/502738: {characterised by integrated valves (throttle valves in microfluidic sample containers B01L3/502746)} - B01L 3/502792: {for moving individual droplets on a plate, e.g. by locally altering surface tension} - B01L 3/502715: {characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces} - B01L 3/5029: {using swabs} - B01L 3/523: {with means for closing or opening} - B01L 3/527: {for a plurality of reagents} - B01L 7/52: {with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples} - B01L 2200/025: Align devices or objects to ensure defined positions relative to each other - B01L 2200/027: for microfluidic devices - B01L 2200/04: Exchange or ejection of cartridges, containers or reservoirs - B01L 2200/0647: Handling flowable solids, e.g. microscopic beads, cells, particles - B01L 2200/141: Preventing contamination, tampering - B01L 2300/044: pierceable, e.g. films, membranes - B01L 2300/0672: Integrated piercing tool - B01L 2300/0681: Filter - B01L 2300/0829: Multi-well plates; Microtitration plates - B01L 2300/0864: comprising only one inlet and multiple receiving wells, e.g. for separation, splitting - B01L 2300/0867: Multiple inlets and one sample wells, e.g. mixing, dilution - B01L 2300/0887: Laminated structure - B01L 2300/089: Virtual walls for guiding liquids - B01L 2300/161: Control and use of surface tension forces, e.g. hydrophobic, hydrophilic - B01L 2400/0427: Electrowetting - B01L 2400/043: magnetic forces - B01L 2400/0478: pistons - B01L 2400/0683: mechanically breaking a wall or membrane within a channel or chamber - G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q) - B01L: CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE ## Abstract A cartridge manipulates samples in liquid droplets with an electrode array when a working film is placed on the array. The cartridge has a body with lower surface and wells to hold samples, each with a bottom opening to release liquid. A piercable bottom structure seals the bottom openings. A working film below the body has a hydrophobic upper surface. A peripheral spacer connects the working film to the body and forms a gap is between the body and surface. A top piercing system located in at least one of the wells has a piston and a piercing element, the piston being movable in the well and the piercing element piercing the piercable bottom structure for releasing a sample from a well into the gap. ## Claims ### Claim 1 A cartridge (1) with a working film (10) and at least one piercing element (13) for manipulating samples in liquid droplets with an electrode array (20) when the working film (10) of the cartridge (1) is placed on said electrode array (20), wherein the cartridge (1) comprises: a) a body (2,2′,2″) that comprises an upper surface (3), a lower surface (4), and a number of wells (5) configured to hold therein reagents (6) or samples (6′), each well (5) comprising a top opening (72), and a bottom opening (73) for releasing a liquid from the well (5); b) a piercable bottom structure (8) impermeable to liquids and configured to seal at least one of the bottom openings (73) of the wells (5); c) a working film (10) located below the lower surface (4) of the body (2,2′,2″), the working film (10) being impermeable to liquids and comprising a hydrophobic upper surface (11); d) a peripheral spacer (9) located below the lower surface (4) of the body (2,2′,2″) and connecting the working film (10) to the body (2,2′,2″); and e) a gap (12) between the lower surface (4) of the body (2,2′,2″) and the hydrophobic upper surface (11) of the working film (10), the gap (12) being defined by the peripheral spacer (9); wherein the cartridge (1) further comprises f) at least one top piercing system (60), each located within at least one of the wells (5) for releasing a reagent or sample (6,6′) from said at least one well (5) into the gap (12); wherein each top piercing system (60) comprises a piston (61) and a piercing element (13), the piston (61) being configured to be movable within said well (5) while providing a seal between the piston (61) and the inner wall of the well (5), and the piercing element (13) being configured as a thorn (63) located at a front side (74) of the piston (61) and being configured to pierce the piercable bottom structure (8) for releasing a reagent or sample (6,6′) from said at least one well (5) into the gap (12) upon moving the piston (61) within the well (5) toward its bottom opening (73). ### Claim 2 (depends on 1) Cartridge (1) according to claim 1, wherein the piercable bottom structure (8) is configured as a piercable foil (79) that is sealingly attached to the lower surface (4) of the body (2,2′,2″). ### Claim 3 (depends on 2) Cartridge (1) according to claim 2, wherein the piercable foil (79) is sized to seal the bottom openings (73) of all wells (5) of the body (2,2′,2″). ### Claim 4 (depends on 1) Cartridge (1) according to claim 1, wherein a front side (74) of the piston (61) of the at least one top piercing system (60) is adapted in shape to a shape of an inner bottom (76) of the well (5) into which the at least one top piercing system (60) is integrated. ### Claim 5 (depends on 4) Cartridge (1) according to claim 4, wherein the inner bottom (76) of the well (5) which comprises the at least one top piercing system (60) is configured as a taper-bottom, and wherein the front side (74) of the piston (61) of said at least one top piercing system (60) is configured as a complementary tapered front side (74). ### Claim 6 (depends on 4) Cartridge (1) according to claim 4, wherein the inner bottom (76) of the well (5), which comprises the at least one top piercing system (60) is configured as a flat bottom, and wherein the front side (74) of the piston (61) of said at least one top piercing system (60) is configured as a complementary flat front side (74). ### Claim 7 (depends on 1) Cartridge (1) according to claim 1, wherein the thorn (63) comprises a neck portion (69) and a tip portion (70) with the tip, the tip portion (70) being configured to pierce the piercable bottom structure (8). ### Claim 8 (depends on 1) Cartridge (1) according to claim 1, wherein the thorn (63) comprises at least one guiding channel (75) for providing an outlet path for releasing the reagent or sample (6,6′) from the well (5) when the thorn (63) of the top piercing system (60) has pierced the piercable bottom structure (8). ### Claim 9 (depends on 7) Cartridge (1) according to claim 7, wherein the tip (70) of the thorn (63) is Phillips head shaped. ### Claim 10 (depends on 1) Cartridge (1) according to claim 1, wherein the top piercing system (60) comprises at least one gasket (62) attached to an upper part (66) of the piston (61) for sealingly abutting the inner wall of the well (5). ### Claim 11 (depends on 10) Cartridge (1) according to claim 10, wherein the at least one gasket (62) is an O-ring, or a quad-ring, or a bellow. ### Claim 12 (depends on 1) Cartridge (1) according to claim 1, wherein the piston (61) and the thorn (63) of the top piercing system (60) are made as a single injection molded piece. ### Claim 13 (depends on 1) Cartridge (1) according to claim 1, wherein the cartridge (1) further comprises a thorn relief spacer layer (64) that is attached to a lower side of the piercable bottom structure (8) and that comprises a relief opening (71) situated below the bottom opening (73) of the well (5), said relief opening (71) providing an additional space for the thorn (63) after piercing the piercable bottom structure (8), and providing an outlet path for releasing the reagent or sample (6,6′) into the gap (12). ### Claim 14 (depends on 13) Cartridge (1) according to claim 13, further comprising a rigid cover (17) that is attached to a lower side of the thorn relief spacer layer (64), said rigid cover (17) enclosing the gap (12) on a side opposite to the working film (10) and comprising at least one cover hole (18) which is situated below the bottom opening (73) of the well (5) and below the relief opening (71) of the thorn relief spacer layer (64). ### Claim 15 (depends on 1) Cartridge (1) of claim 1, wherein the peripheral spacer (9) surrounds a part or the entire foot print of the cartridge (1) and is attached to the lower surface (4) of the piercable bottom structure (8). ### Claim 16 (depends on 15) Cartridge of claim 15, wherein the cartridge (1) comprises at least one intermediate spacer (15) configured reaching from the peripheral spacer (9) of one side of the cartridge (1) to the peripheral spacer (9) of another side of the cartridge (1) or as a separate element located within the area of the gap (12), the intermediate spacer (15) being attached to the lower surface (4) of the piercable bottom structure (8). ### Claim 17 (depends on 1) The cartridge of claim 1, wherein the working film (10) is configured as a monolayer of a hydrophobic material. ### Claim 18 (depends on 1) The cartridge of claim 1, wherein the working film (10) is configured as a monolayer of electrically non-conductive material, the upper surface (11) of the working film (10) being treated to be hydrophobic. ### Claim 19 A system (40) for liquid droplet manipulation, the system comprising a substrate (42) with an electrode array (20) and a central control unit (43) for controlling the selection of individual electrodes (44) of the electrode array (20) and for providing the electrodes (44) with individual voltage pulses for manipulating liquid droplets (23) by electrowetting, wherein the system (40) is configured to receive on top of the electrodes (44) a cartridge (1) with a working film (10) and at least one piercing element (13) for manipulating samples in liquid droplets with an electrode array (20) when the working film (10) of the cartridge (1) is placed on said electrode array (20), wherein the cartridge (1) comprises: a) a body (2,2′,2″) that comprises an upper surface (3), a lower surface (4), and a number of wells (5) configured to hold therein reagents (6) or samples (6′), each well (5) comprising a top opening (72), and a bottom opening (73) for releasing a liquid from the well (5); b) a piercable bottom structure (8) impermeable to liquids and configured to seal at least one of the bottom openings (73) of the wells (5); c) a working film (10) located below the lower surface (4) of the body (2,2′,2″), the working film (10) being impermeable to liquids and comprising a hydrophobic upper surface (11); d) a peripheral spacer (9) located below the lower surface (4) of the body (2,2′,2″) and connecting the working film (10) to the body (2,2′,2″); and e) a gap (12) between the lower surface (4) of the body (2,2′,2″) and the hydrophobic upper surface (11) of the working film (10), the gap (12) being defined by the peripheral spacer (9); wherein the cartridge (1) further comprises f) at least one top piercing system (60), each located within at least one of the wells (5) for releasing a reagent or sample (6,6′) from said at least one well (5) into the clap (12); wherein each top piercing system (60) comprises a piston (61) and a piercing element (13), the piston (61) being configured to be movable within said well (5) while providing a seal between the piston (61) and the inner wall of the well (5), and the piercing element (13) being configured as a thorn (63) located at a front side (74) of the piston (61) and being configured to pierce the piercable bottom structure (8) for releasing a reagent or sample (6,6′) from said at least one well (5) into the gap (12) upon moving the piston (61) within the well (5) toward its bottom opening (73). ### Claim 20 (depends on 19) The system of claim 19, wherein the system (40) comprises actuating elements (41) for actuating the piston (61) of at least one top piercing system (60) of a cartridge (1) for releasing reagents, treatment liquids, oil, reaction liquids or sample containing liquids into the gap (1 2) of the cartridge (1). ### Claim 21 (depends on 19) The system of claim 19, wherein the substrate (42) comprises an electrically insulating film, layer or cover (50) that is applied to the electrode array (20), that covers all individual electrodes (44) of the electrode array (20) and that separates the individual electrodes (44) from each other. ### Claim 22 A method of releasing a reagent or sample (6,6′) from a well (5) of a cartridge (1) with a working film (10) and at least one piercing element (13) for manipulating samples in liquid droplets with an electrode array (20) when the working film (10) of the cartridge (1) is placed on said electrode array (20), wherein the method comprises the following steps: providing a cartridge (1) which comprises: a) a body (2,2′,2″) that comprises an upper surface (3), a lower surface (4), and a number of wells (5) configured to hold therein reagents (6) or samples (6′), each well (5) comprising a top opening (72), and a bottom opening (73) for releasing a liquid from the well (5); b) a piercable bottom structure (8) impermeable to liquids and configured to seal at least one of the bottom openings (73) of the wells (5); c) a working film (10) located below the lower surface (4) of the body (2,2′,2″), the working film (10) being impermeable to liquids and comprising a hydrophobic upper surface (11); d) a peripheral spacer (9) located below the lower surface (4) of the body (2,2′,2″) and connecting the working film (10) to the body (2,2′,2″); e) a gap (12) between the lower surface (4) of the body (2,2′,2″) and the hydrophobic upper surface (11) of the working film (10), the gap (12) being defined by the peripheral spacer (9); and f) at least one top piercing system (60), each located within at least one of the wells (5) for releasing a reagent or sample (6,6′) from said at least one well (5) into the gap (12); wherein each top piercing system (60) comprises a piston (61) and a piercing element (13), the piercing element (13) being configured as a thorn (63) located at a front side (74) of the piston (61); moving the piston (61) within said well (5) toward its bottom opening (73) while providing a seal between the piston (61) and the inner wall of the well (5); piercing the piercable bottom structure (8) with the thorn (63); and releasing a reagent or sample (6,6′) from said at least one well (5) into the gap (12) upon moving the piston (61) within the well (5). ### Claim 23 A system configured to perform a method of releasing a reagent or sample (6,6′) from a well (5) of a cartridge (1) with a working film (10) and at least one piercing element (13) for manipulating samples in liquid droplets with an electrode array (20) when the working film (10) of the cartridge (1) is placed on said electrode array (20), wherein the method comprises the following steps: providing a cartridge (1) which comprises: a) a body (2,2′,2″) that comprises an upper surface (3), a lower surface (4), and a number of wells (5) configured to hold therein reagents (6) or samples (6′), each well (5) comprising a top opening (72), and a bottom opening (73) for releasing a liquid from the well (5); b) a piercable bottom structure (8) impermeable to liquids and configured to seal at least one of the bottom openings (73) of the wells (5); c) a working film (10) located below the lower surface (4) of the body (2,2′,2″), the working film (10) being impermeable to liquids and comprising a hydrophobic upper surface (11); d) a peripheral spacer (9) located below the lower surface (4) of the body (2,2′,2″) and connecting the working film (10) to the body (2,2′,2″); e) a gap (12) between the lower surface (4) of the body (2,2′,2″) and the hydrophobic upper surface (11) of the working film (10), the gap (12) being defined by the peripheral spacer (9); and f) at least one top piercing system (60), each located within at least one of the wells (5) for releasing a reagent or sample (6,6′) from said at least one well (5) into the gap (12); wherein each top piercing system (60) comprises a piston (61) and a piercing element (13), the piercing element (13) being configured as a thorn (63) located at a front side (74) of the piston (61); moving the piston (61) within said well (5) toward its bottom opening (73) while providing a seal between the piston (61) and the inner wall of the well (5); piercing the piercable bottom structure (8) with the thorn (63); and releasing a reagent or sample (6,6′) from said at least one well (5) into the gap (12) upon moving the piston (61) within the well (5).