Microvolume liquid dispensing array

[0002] This application claims priority from U.S. provisional application Ser. No. 60/185,810, filed Feb. 29, 2000.

[0003] This invention relates to microfluidics and laboratory automation.

[0004] The development of automated combinatorial chemistry systems and ultra-high throughput screening systems have dramatically increased the number of compounds per unit time being synthesized and screened in drug discovery programs. Such technology involves rapid handling of large numbers of very small samples. For example, thousands of new compounds per week may be produced, with each compound being concentrated in a total volume of only 50 microliters. Microliter amounts of sample often must suffice for hundreds of screening assays. Conventionally, aliquots of the concentrated, liquid sample are dispensed using “sip and spit” liquid handling technology, diluted in an appropriate medium, and re-dispensed into an assay mixture, again using sip and spit technology. This “reformatting” process adds complexity to the overall process, thereby increasing time and cost per assay. In addition, reformatting generates waste of valuable sample material.

[0005] The invention features a method of packaging a multiplicity of liquids for shipment, storage and metered dispensing. The method includes: (a) providing an integrated array of isolated reservoir units alignable with an array of liquid-receiving units (LRUs); (b) dispensing the liquids into the array of reservoir units; and (c) incorporating a dispensing tap into each reservoir unit to form a reservoir/tap unit sealed against spillage or leakage of the liquids. Preferably, the reservoir units are also sealed against air and light. The array of LRUs can be a multiwell container such as a 96-well microtiter plate, a 384-well microtiter plate, or a 1536-well microtiter plate. In preferred embodiments, each tap includes a translatable metering tube, which contains a tube end closure, a port, and a translatable piston. In some embodiments, the liquid is a solution of one or more chemical compounds. In some embodiments, liquid-contacting surfaces of the reservoir and tap are resistant to damage by acids, bases, salts and organic solvents.

[0006] The invention also features a method for independently dispensing a metered amount of a plurality of liquids into an array of LRUs. The method includes: (a) providing an array of isolated, sealed, tapped reservoir units, the array of reservoir units including a reservoir for each LRU, each reservoir unit containing an integrated metering tap; (b) aligning the array of reservoir/tap units with the array of LRUs so that each tap is aligned with one LRU; and (c) actuating one or more taps in the array of reservoir units so that each actuated tap dispenses a metered amount of liquid into the LRU aligned with that tap. The metered amount dispensed into any particular unit in the array can be from zero nanoliters to 20 microliters, preferably from 20 nanoliters to 2 microliters, e.g., 50 nanoliters to 500 nanoliters.

[0007] Preferably no tap contacts an LRU surface, and the liquid dispensed from each tap breaks contact with the tap before contacting the LRU aligned with that tap or the contents of an LRU. Preferably, the reservoirs are sealed against air and light. The array of reservoir units can be aligned directly above the array of LRUs. In some embodiments, each tap can be actuated independently. Preferably, each tap contains minimal (or substantially zero) dead volume. Examples of suitable LRUs are multi-well containers such as a 96-well microtiter plate, a 384-well microliter plate and a 1536-well microtiter plate.

[0008] In some embodiments of the dispensing method, each tap includes a translatable metering tube, which can contain a tube end closure, a port and a translatable piston. Actuating the tap can include translating the tube so that the port is inside the reservoir; drawing liquid from the reservoir through the port and into the tube; translating the tube so that the port is outside the reservoir; and expelling liquid from the tube through the port and into a fluid output channel. The liquid can be drawn into the tube and expelled from the tube by translating the piston. Some embodiments include propelling the expelled liquid away from the port. Propelling the expelled liquid can be achieved by applying a propelling fluid to the expelled liquid. The propelling fluid can be a propelling liquid, e.g., an aqueous liquid or an organic solvent; or a propelling gas, e.g., air, nitrogen or argon. Some embodiments of the method include providing a curtain of forced gas surrounding the fluid output tip, with the forced gas moving in the same direction as the liquid exiting from the fluid output tip.

[0009] The invention also features devices for storing, shipping and dispensing metered, nanoliter or microliter amounts of liquid into a liquid receiving unit.

[0010] An offset nozzle-type device includes: an array of isolated, sealed, reservoir/tap units, each unit containing an integrated metering tap, each tap including: (a) a metering tube translatable between a fill position inside the reservoir and an expel position outside the reservoir. The metering tube includes (1) a tube end closure, e.g., a plug, in a lower portion of the tube, (2) a port above the tube end closure, and (3) a piston in an upper portion of the tube. The piston is movable between a down position that seals the port and an up position above the port; and (b) a fluid output channel having an upper portion in fluid communication with the port when the tube is in the expel position and a lower portion terminating in a fluid output tip. A compressed gas path in fluid communication with the fluid output channel at a point upstream of the port when the tube is in the expel position can be used to apply a gas stream to propel the expelled liquid through the fluid output channel. Some embodiments include a compressed gas path terminating in an annular opening surrounding the fluid output tip.

[0011] An in-line nozzle embodiment of the device includes an array of isolated, sealed reservoir/tap units, each unit containing an integrated metering tap, each tap including: (a) a metering tube translatable between a fill position inside the reservoir and an expel position outside the reservoir. The metering tube contains (1) a tube end closure in a lower portion of the tube, (2) a port above the tube end closure, and (3) a piston in an upper portion of the tube. The piston is movable between a down position that seals the port and an up position above the port; and (b) a nozzle containing a fluid output channel through which the tube extends when in the down position, the fluid output channel having an upper end in fluid communication with a compressed gas path, and a lower end terminating in a nozzle tip.

[0012] A nozzleless-type device includes an array of isolated, sealed reservoir/tap units, each unit containing an integrated metering tap. Each metering tap including a metering tube translatable between a fill position inside the reservoir and an expel position outside the reservoir. The metering tube contains (1) a tube end closure in a lower portion of the tube, (2) a port above the tube end closure, and (3) a piston in an upper portion of the tube. The piston is movable between a down position that seals the port and an up position above the port. Each unit contains a compressed gas path, which includes one or more compressed gas outlets located above the port so that it can deliver a downward gas stream across the port, when the metering tube is in the expel position.

[0013] In each of the above devices, movement of the piston from the up position to the down position can displace, for example, 10 nanoliters to 20 microliters, preferably from 20 nanoliters to 2 microliters, e.g., 50 nanoliters to 500 nanoliters. The array of reservoir units can be arranged so that each tap aligns with one well of a multi-well container such as a 96-well microtiter plate, a 384-well microtiter plate or a 1536-well microtiter plate. However, with suitable equipment, any particular tap can be positioned to dispense into any chosen well.

[0014] As used herein, “liquid-receiving unit” (LRU) means: (a) a defined or addressable area on a flat liquid-receiving surface, e.g., a glass slide; (b) a depression or well in a liquid-receiving container, e.g., a microtiter plate, or (c) a receptacle, e.g., a test tube, vial or bottle.

[0015] As used herein, “reservoir/tap unit” means a single tapped reservoir.

[0016] The details of one or more embodiments of the invention are set forth in the accompanying drawings and the description below. Other features, objects, and advantages of the invention will be apparent from the description and drawings, and from the claims.

[0017] FIG. 1 is a perspective view of a device for integrated storage and single-channel dispensing of small volumes of liquids. The depicted device contains a 96-unit array of integrated reservoir/tap units. The 96 units are arranged so that each of the 96 tips aligns with one well of a conventional 96-well microtiter plate.

[0018] FIG. 2 is a sectional view of a single reservoir/tap unit. The unit has a metering tube which is in the up position. FIG. 2 depicts an offset nozzle embodiment.

[0019] FIG. 3 is a sectional view of the reservoir/tap unit shown in FIG. 2, but with the metering tube in the down position.

[0020] FIG. 4 is a detail from FIG.2. The enlarged detail view shows the tube in the up position, and a piston in the tube. The piston is in the down position, where it rests against a tube plug.

[0021] FIG. 5 is the same as FIG. 4, except that the piston is raised into an up position.

[0022] FIG. 6 is an enlarged, front view of a metering tube. A tube port, through which liquid enters and leaves the tube is visible near the lower end of the tube.

[0023] FIG. 7 is a detail enlarged from FIG. 6, showing the tube port.

[0024] FIG. 8 is an enlarged sectional view (rotated 90° relative to FIG. 6) showing a lower portion of the tube, the tube plug, and a lower portion of the piston.

[0025] FIG. 9 is a sectional view (detail) of a reservoir/tap unit in which the metering tube is in the down position, and the piston is in an up position.

[0026] FIG. 10 is a sectional view (detail) of a reservoir/tap unit in which the metering tube is in the down position, and the piston is in a down position.

[0027] FIG. 11 is the same as FIG. 10, except that it shows a metered amount of liquid in an upper region of a fluid flow path, and arrows indicating flow of compressed gas through a compressed gas path.

[0028] FIG. 12 is the same as FIG. 11, except that it shows the metered amount of liquid in a middle region of the fluid flow path, and arrows indicating flow of compressed gas sweeping the liquid down the fluid flow path.

[0029] FIG. 13 is the same as FIG. 12, except that it shows the liquid in the lowermost portion of the fluid flow path, where the liquid is exiting from a flow path tip.

[0030] FIG. 14 is a sectional view of a single reservoir/tap unit. The unit has a metering tube in the up position. FIG. 14 depicts an in-line nozzle embodiment.

[0031] FIG. 15 is a sectional view of the reservoir/tap unit shown in FIG. 14, but with the metering tube in the down position.

[0032] FIG. 16 is a detail from FIG.14. The enlarged detail view shows the tube in the up position, and a piston in the tube. The piston is in the down position, where it rests against a tube plug.

[0033] FIG. 17 is the same as FIG. 16, except that the piston is raised into an up position.

[0034] FIG. 18 is a sectional view (detail) of a reservoir/tap unit in which the metering tube is in the down position, and the piston is in an up position.

[0035] FIG. 19 is a sectional view (detail) of a reservoir/tap unit in which the metering tube is in the down position, and the piston is in the down position. FIG. 19 shows a bolus of expelled liquid emerging from a port in the side of the tube.

[0036] FIG. 20 is a sectional view (detail) of a reservoir/tap unit (in-line nozzle) in which the metering tube has been withdrawn from the down position to the up position, after expulsion of a bolus of liquid. The liquid has been drawn into the fluid output channel in the nozzle.

[0037] FIG. 21 is a sectional view of a single nozzleless tapped reservoir unit with the metering tube in the up position.

[0038] FIG. 22 is a sectional view of a nozzleless reservoir/tap unit with the metering tube in the down position. A bolus of expelled liquid is shown at the port.

[0039] FIG. 23 is a sectional view of a nozzleless reservoir/tap unit with the metering tube in the down position. A bolus of expelled liquid is shown in flight after being propelled from the tip of the metering tube.

[0040] FIG. 24 is a detail from FIG. 21. The enlarged detail view shows the metering tube in the up position and the piston in the down position.

[0041] FIG. 25 is the same as FIG. 24, except that the metering tube is in the up position and the piston is in the up position.

[0042] FIG. 26 is the same as FIG. 24, except that the metering tube is in the down position and the piston is in the up position.

[0043] FIG. 27 is the same as FIG. 24, except that the metering tube is in the down position and the piston is in the down position.

[0044] FIGS. 28A-28F are sectional views depicting a device and sequence of events in a preferred packaging method.

[0045] Throughout the various drawings, like reference numbers indicate like elements.

[0046] The invention provides methods and devices for integrated packaging, shipping, storage, and dispensing of extremely small volumes of liquids, e.g., aqueous solutions and compounds dissolved in organic solvents, in an automated, multi-well format of the type used in high throughput screening (HTS) or ultra-high throughput screening (UHTS). By virtue of a metering tap integrated with each reservoir in an array of reservoir/tap units, the invention advantageously avoids the use of conventional sip and spit technology. Consequently, multi-well plate assays can be performed without reformatting, i.e., transferring aliquots of concentrated samples from storage plates to working plates, diluting on working plates, transferring diluted samples from working plates to assay plates, etc. This maximizes speed and efficiency. Entire sets of samples, e.g., compounds for screening, can be stored and/or shipped conveniently in a single cassette, which can be plugged into an HTS or UHTS system, where nanoliter volumes of concentrated sample can be dispensed directly onto assay plates without reformatting. Because the reservoir/tap units in an array are isolated from each other, single-channel dispensing is achieved, and each reservoir/tap unit is individually addressable. Because each reservoir/tap unit in an array (cassette) can be sealed against air, moisture and light, labile compounds can be stored and handled under favorable conditions.

[0047] FIG. 1 is a perspective view of a device 10 according to the invention for storing and dispensing liquid into a conventional 96-well microtiter plate 12. Protruding from lower surface 13 of device 10 are 96 flow tips 11 arranged so that when device 10 is aligned above 96-well microtiter plate 12, each tip 11 is above a different one of the 96 wells 14 in plate 12. On the upper surface 15 of device 10 are 96 mechanical interfaces 16 for tap actuation. Operation of each interface 16 actuates a tap whose flow path tip 11 is located beneath that interface 16.

[0048] FIG. 2 is a sectional view of a single reservoir/tap unit 20. The unit 20 contains a reservoir 21 formed by a cylinder wall 22, sliding seal 23 and lower seal 24. The unit 20 also contains a metering tube 25, tube handle 26, tube handle spring 27, piston handle 28, piston 29, and piston handle stop 30. The embodiment depicted in FIG. 2 is an example of an offset nozzle embodiment, because nozzle tip 11 is not directly in line with metering tube 25. FIG. 2 shows the tube 25 and tube handle 26 in the up position. Tube handle 26 and piston handle 28 are included in each mechanical interface 16 shown in FIG. 1.

[0049] Reservoir 21 contains minimal air space. Therefore, liquid in reservoir 21 is essentially coextensive with the volume of reservoir 21. As liquid is metered from reservoir 21, sliding seal 23 slides downward reducing the volume of reservoir 21 so that remnant liquid in reservoir 21 remains coextensive with the volume of reservoir 21. FIG. 3 is a sectional view of a single unit 20 in which the tube 25 is in the down position.

[0050] FIG. 4 is a detail from FIG. 2, in which tube 25 is in the up position and piston 29 is in the down position. In the down position, piston 29 rests against tube plug (tube end closure) 31 so that piston 29 closes and seals tube port 32, thereby blocking entry of liquid from reservoir 21 into tube 25. FIG. 5 is the same as FIG. 4, except that piston 29 is raised into an up position. Raising piston 29 opens tube port 32 and draws a metered amount of liquid from reservoir 21 into tube 25, with the metered amount depending on the height to which piston 29 is raised.

[0051] FIG. 6 is an enlarged, front view of metering tube 25, showing tube port 32. FIG. 7 is a detail from FIG. 6, showing tube port 32. FIG. 8 is an enlarged sectional view (rotated 90° relative to FIG. 6) showing a lower portion of tube 25, tube plug 31, and a lower portion of piston 29.

[0052] In illustrating operation of device 20, FIG. 9 is sequential, following FIG. 5. In FIG. 9, metering tube 25 has been translated downward into the down position, with piston 29 remaining in the up position, i.e., same position relative to tube 25. In FIG. 9, downward translation of tube 25 through lower seal 24 has taken port 32 out of reservoir 21 and placed port 32 in fluid communication with fluid output channel 33. The next sequential step is lowering of piston 29 into the down position, in which piston 29 rests against tube plug 31. This lowering of pistion 29 expels liquid (not shown) from tube 25 and into fluid output path 33.

[0053] FIG. 11 is the same as FIG. 10, except that it shows the expelled liquid 40 in an upper region of fluid output channel 33, and arrows A indicating flow of compressed air through a compressed gas path 34, and exit of the compressed air from an annular compressed gas outlet 35 surrounding nozzle tip 11. The exiting air forms an annular curtain of air moving downward and surrounding a droplet of liquid that will exit from nozzle tip 11. The annular curtain of air facilitates controlled movement of the droplet into the correct well, and effectively isolates all droplets and corresponding wells from each other.

[0054] FIG. 12 is the same as FIG. 10, except that it shows a metered amount of expelled liquid 40 (bolus), in a middle region of fluid output channel 33, and arrows B indicating flow of compressed air in fluid output channel 33. Air flowing from compressed gas inlet 59 through fluid output channel 33 sweeps liquid 40 down fluid output channel 33. FIG. 13 shows liquid 40 in the lowermost portion of fluid output channel 33, where it is exiting nozzle tip 11.

[0055] FIG. 14 is a sectional view of a single reservoir/tap unit 20. The unit 20 contains a reservoir 21 formed by a cylinder wall 22, and lower seal 24. The unit 20 also contains a metering tube 25, tube handle 26, piston handle 28, piston 29, and piston handle stop 30. The embodiment depicted in FIG. 14 is an example of an in-line nozzle embodiment, because nozzle tip 11 is directly in line with metering tube 25. FIG. 14 shows the tube 25 and tube handle 26 in the up position. Tube handle 26 and piston handle 28 are included in each mechanical interface 16 shown in FIG. 1. FIG. 15 is a sectional view corresponding to FIG. 14, except that tube 25 is in the down position.

[0056] FIG. 16 is a detail from FIG. 14, in which tube 25 is in the up position and piston 29 is in the down position. In the down position, piston 29 rests against tube plug 31 so that piston 29 closes and seals tube port 32, thereby blocking entry of liquid from reservoir 21 into tube 25. FIG. 17 is the same as FIG. 16, except that piston 29 is raised into an up position. Raising piston 29 opens tube port 32 and draws a metered amount of liquid from reservoir 21 into tube 25, with the metered amount depending on the height to which piston 29 is raised.

[0057] In FIG. 18, metering tube 25 has been translated downward into the down position, with piston 29 remaining in the up position, i.e., same position relative to tube 25. In this in-line nozzle embodiment of the invention, tube 25 passes through fluid output channel 33 as it translates between the up position and the down position. In the down position, port 32 is beneath nozzle tip 11. The next sequential step is lowering of piston 29 into the down position, in which piston 29 rests against tube plug 31, as shown in FIG. 19. This lowering of piston 29 expels liquid from tube 25 through port 32.

[0058] A bolus of expelled liquid 40 is shown in FIG. 19. Expelled liquid 40 clings to the side of tube 25 as a result of surface tension and adhesion. As tube 25 is retracted, nozzle tip 11 forces expelled liquid 40 to slide down the outside of tube 25. During retraction, when bottom end 42 of tube 25 reaches lower end 44 of fluid output channel 33, expelled liquid 40 migrates to bottom end 42 of tube 25 and clings there. As tube 25 is further retracted, expelled liquid 40 follows bottom end 42 of tube 25 upward through fluid output channel 33 (FIG. 20). When bottom end 42 of tube 25 reaches upper end 43 of fluid output channel 33, expelled liquid 40 detaches from bottom end 42 of tube 25 and remains in upper portion of fluid output channel 33. When tube 25 is fully retracted into up position, compressed air enters compressed gas path 34 and pushes expelled liquid 40 downward, so that it exits nozzle tip 11 and falls into a well in a microtiter plate (not shown).

[0059] In in-line nozzle embodiments of the invention, nozzle 45 preferably is made of an elastomeric material, with fluid output channel 33 having an inside diameter slightly smaller than the outside diameter of tube 25. Fluid output channel 33 expands slightly to accommodate tube 25, as the tube passes through the fluid output channel. This promotes an airtight seal between tube 25 and fluid output channel 33, when the tube is in the channel. Selection of a suitable elastomer is within ordinary skill in the art.

[0060] FIGS. 21-27 depict a nozzleless tapped reservoir. In FIG. 21 reservoir/tap unit 20 has metering tube 25 in the up position and piston 29 in the down position. FIG. 24 is a detail from FIG. 21 in which tube 25 is in the up position and piston 29 is in the down position. In the down position, piston 29 rests against tube plug (tube end closure) 31 so that piston 29 closes and seals tube port 32, thereby blocking entry of liquid 40 from reservoir 21. FIG. 25 is the same as FIG. 24, except that piston 29 is raised into an up position. Raising piston 29 opens tube port 32 and draws a metered amount of liquid from reservoir 21 into tube 25, with the metered amount depending on the height to which piston 29 is raised. In FIG. 26, metering tube 25 has been translated downward into the down position, with piston 29 remaining in the up position. In this nozzleless embodiment of the invention, no fluid output channel or nozzle is necessary, and port 32 and fine point 46 are exposed (FIG. 22). The next sequential step is lowering of piston 29 into the down position, where it rests against tube plug 31, (FIG. 27). This expels a precisely metered amount of liquid 40 through port 32. Expelled liquid 40 is then swept downward by a downward flow of air from compressed gas outlet 35. FIG. 23 shows expelled liquid 40 dropping from bottom end 42 of metering tube 25, which is tapered to a fine point 46. Fine point 46 facilitates release of expelled liquid 40 from bottom end 42 of metering tube 25 in a controlled manner. In some embodiments of the invention, a shroud 57 surrounds or partially shields lower end 48 of metering tube 25, which extends downward when tube 25 is in the expel position.

[0061] FIGS. 28A-28F depict a device and sequence of events in a preferred packaging method according to the invention. Two reservoir/tap units in an array are depicted. In FIG. 28A fill pin 51, which has an outside diameter equal to that of metering tube 25, extends upward through lower seal 24. This permits dispensing of liquid 40 into reservoir 21 from reservoir filling device 52 positioned above the reservoir. In FIG. 28B cap seal 53 is installed on top of reservoir 21. In FIG. 28C metering tube 25 is aligned directly above fill pin 51, so that tube bottom end 42 contacts upper end 54 of fill pin 51. In FIG. 28D metering tube 25 is lowered so as to push fill pin 51 downward. By this process, metering tube 25 replaces fill pin 51 without allowing leakage of liquid 40 from reservoir 21. In FIG. 28E metering tube 25 is seated against lower seal 24. In FIG. 28F protective cover 55 is installed on bottom of device 20 for storage. In FIG. 28E, it can be seen that the lower portion of the device serves as a shroud 49 around lower end 48 of metering tube 25.

[0062] Devices according to the invention can be designed for compatibility with various liquids, including aqueous buffers, organic solvents, e.g., dimethylsulfoxide, acids and bases. Compatibility is achieved by selection of suitable materials for fabrication of components that contact the liquid. Exemplary materials for fabrication of components are stainless steel, nylon, polyethylene, polypropylene, EPD rubber and polytetrafluoroethylene (PTFE; Teflon®). Selection of suitable materials and fabrication of components is within ordinary skill in the art.

[0063] It is to be understood that various modifications on the above-described embodiments can be made without departing from the spirit and scope of the invention. For example, to form a liquid reservoir, sliding seal 23 and lower seal 24 can be replaced with an expandable bladder. Accordingly, other embodiments of the invention are within the scope of the following claims.