WO2007022667A1

WO2007022667A1 - Multiple autopipette apparatus and method of operation

Info

Publication number: WO2007022667A1
Application number: PCT/CN2006/000014
Authority: WO
Inventors: Dong Wang; Jingchun Zhang; Wei Gao; Yingna Li; Kun Zou; Jing Cheng
Original assignee: Capitalbio Corporation; Tsinghua University
Priority date: 2005-08-26
Filing date: 2006-01-06
Publication date: 2007-03-01
Also published as: CN1736611A; CN100402153C

Abstract

An embodiment of the present invention comprises an automatic multichannel pipettor operating in coordination with a robotic positioner. A volumetrically apportioned pressure dividing manifold equally divides volumetric displacement from a syringe pump among a plurality of pipetting channels for simultaneous aspiration and dispensing. The automatic pipettor can use disposable pipette tips that can be automatically loaded and discarded.

Description

MULTIPLE AUTOPIPETTE APPARATUS AND METHOD OF OPERATION

Technical Field

[0001] The present invention relates to an automated pipetting apparatus ("autopipettor") for the simultaneous delivery of multiple, separate aliquots of sample fluids.

Background Art

[0002] Pipetting systems are widely used for clinical, laboratory, and manufacturing applications. There are two categories of pipetting system, manual and automatic. Manual pipette systems can be actuated by breath pressure, or by manipulation of an elastomeric bulb or a piston pump to aspirate and dispense fluid. Automatic pipette systems often use a motor driven syringe pump to aspirate, and dispense a metered volume of sample fluid. Automatic pipette systems can be combined with robotic positioning technology, for coordinated positioning, aspiration, and fluid dispensing. Applications for automatic pipetting include, without exclusion, sample treatment, sample purification, sample amplification, electrophoresis, liquid chromatography, flow cytometry, microarray analysis, and many others. Pipetting technology is indispensable for life sciences fields such as genomics and proteomics. Pipetting can be used to mix, as well as to transfer fluids.

[0003] Multichannel autopipettors can simultaneously transfer, for example, 4, 8, 12, 24, or 96 aliquots of fluid samples. Methods of metering volumes of dispensed fluids include volumetric and time methods. For the volumetric method, the volume displacement of the syringe pump is used to estimate the volume of the aspirated or dispensed fluid. The time method estimates the dispensed volume of fluid based on the time interval between opening and closing a valve controlling the fluid discharge combined with known fluid flow rates.

[0004] Currently, multichannel autopipettors based on volumetric methods generally conform to one of two basic configurations. In a first configuration, a pump body comprises multiple syringe cavities, each cavity being coupled to its own pipette tip. The corresponding syringe plungers are mechanically coupled to one another and are driven by a common actuation mechanism. The drawbacks of the first configuration include the need to maintain tight mechanical tolerances in a multiplicity of syringe cavities in a common pump body, as well as the possible need to discard an entire pump body in the event that a single syringe pump fails. Also, volumetric delivery consistency among the different syringe pumps can be difficult to maintain.

[0005] A second basic configuration simply gangs a number of individual syringe pumps, each coupled to its corresponding pipette tip, and each syringe pump having its own, independent actuator. Tecan Corp. of Switzerland adopts this type of configuration. Although the independent actuation of each syringe pump admits the possibility of simultaneously dispensing aliquots of different volumes, the redundant actuator hardware increases equipment costs.

[0006] What is needed is an automatic multichannel pipettor, having a single syringe pump with a single actuator that can simultaneously dispense aliquots of consistent volume through its channels.

Summary of the Invention

[0007] An embodiment of the present invention comprises an automatic multichannel pipettor operating in coordination with a robotic positioner. A volumetrically apportioned, pressure dividing manifold equally divides volumetric displacement from a syringe pump among a plurality of pipetting channels for simultaneous aspiration and dispensing. The automatic pipettor can use disposable pipette tips that can be automatically loaded and discarded.

Brief Description of the Drawings

[0008] Fig. 1 is a schematic drawing of an embodiment of the invention.

[0009] Fig. 2 is a perspective view of an embodiment of the invention.

[0010] Fig. 3 is a cross-sectional view of a detail of an embodiment of the invention.

[0011] Fig. 4 illustrates an operation of an embodiment of the invention.

[0012] Fig. 5 illustrates a further operation of an embodiment of the invention. [0013] Fig.6 illustrates another operation of an embodiment of the invention. [0014] Fig. 7 illustrates a block diagram of a controller for an embodiment of the invention.

Detailed Description of Embodiments of the Invention

[0015] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as is commonly understood by one of ordinary skill in the art to which this invention belongs. AU patents, applications, published applications and other publications referred to herein are incorporated by reference in their entirety. If a definition set forth in this section is contrary to or otherwise inconsistent with a definition set forth in applications, published applications and other publications that are herein incorporated by reference, the definition set forth in this section prevails over the definition that is incorporated herein by reference.

[0016] As used herein, "a" or "an" means "at least one" or "one or more."

[0017] Similar numerical references refer to similar features within the various drawings.

[0018] Fig. 1 is a schematic illustration of an embodiment of the invention having four pipetting channels. Syringe pump 103 is connected to inlet port 10 Ie of the volumetrically apportioned pressure dividing manifold ("manifold") 101 via coupling 108. The plunger of syringe pump 103 is mechanically coupled to actuator 103b through a mechanical coupling 103a. Under the control of controller 110, actuator 103b causes syringe pump 103 to aspirate or expel volumes of air or other working fluid or gas contained therein. Actuator 103b is typically electromechanical, and is coupled to the piston of syringe pump 103 via a mechanical coupling 103 a. Examples of actuator 103 d can include stepper motors and various servo motors, coupled with gears and/or a worm drive for precise control of the position of the piston within syringe pump 103.

[0019] Manifold 101 is shown dividing the pressure or vacuum generated by the syringe pump 103 among four different pipette tips 102a, 102b, 102c, and 102d. It is important that the volume displacement of the syringe pump 103 is equally apportioned between outlets 101a, 101b, 101c, and 1 Old of the manifold so that aliquots of equal volume may be simultaneously dispensed by each pipette tip. Although manifold 101 has four outlets in the embodiment of Fig. 1, in general any number of outlets may be desired. The bore diameters of outlets 101a, 101b, 101c, and 101d can be individually set to compensate for pressure gradients along distributing channel 101f, regardless of the position of the outlet on the manifold. For example, the bore diameters of outlets 101a and 101d, can be larger than the bore diameters of outlets 101b and 101c, to equalize the apportionment of volumetric displacement from syringe pump 103. Rather than, or in addition to, customizing the bore diameters of the outlets, distributing channel 101f can be tapered in cross-section to equalize the apportionment of volumetric displacement on other embodiments of the invention. In further embodiments, buffer zones may be disposed between adjacent outlets to equalize the apportionment of volumetric displacement, as described below.

[0020] Referring again to Fig. 1, pipette tip remover 106 can be a bar with holes therein to accommodate each outlet of the manifold. Pipette tips 102a, 102b, 102c, and 102d are typically made of somewhat mechanically compliant polymer material and are removably press fit onto outlets 101a, 101b, 101c, and 101d, respectively. When pipette tip ejector 106 is actuated downward, as shown by the arrows, pipette tips 102a, 102b, 102c, and 102d are ejected from their holders. Actuator 106b for pipette tip ejection can be electromechanical or pneumatic.

[0021] In an embodiment, the tip ejector actuator can be a pneumatic cylinder. The cylinder can be a double acting cylinder or single acting cylinder. The double acting cylinder has two input ports for the discharged air, which are used to push out and pull in the cylinder shaft independently; the port through which the discharged air is input into the cylinder is controlled through an electric controlled 2-way and 5-port valve; the 2- way and 5-port valve has one air inlet, two air outlets and two air discharging ports; a normally closed 2-way direct operated valve is provided between the air inlet of the 2- way and 5-port valve and the source of the discharged air. The cylinder shaft is fixed on the remover to unload disposable tip on which the through holes are bored which can let the disposable tip holders pass through. The diameter of the through holes is larger than the outer diameter of a tip holder but smaller than the largest diameter of a disposable tip.

[0022] When a disposable tip is to be unloaded, the airway between the inlet of the 2- way and 5-port valve and one of its outlets is open, while the said outlet is connected with the inlet of the cylinder which is used to push out the cylinder shaft, then the 2-way direct operated valve is opened. Under the effect of the discharged air, the cylinder shaft is pushed out, and the ejector fixed with the cylinder shaft to unload the disposable tip moves downward, so that each disposable tip is pushed down from its holder. After that, under the control of controller 110, the airway of the valve's air inlet and its outlet which connects with air inlet of the cylinder that is used to pull in the cylinder shaft is connected. Under the effect of the discharged air, the cylinder shaft is pulled in; then the 2-way direct operated valve is changed back to its normally closed position, and the operation to unload disposable tip is finished.

[0023] In various embodiments of the invention, the manifold and the disposable tip holder can be formed integrally, or they may be formed separately and assembled together. In further embodiments, a non-disposable tip can replace the combination of the disposable tip and the disposable tip holder.

[0024] Fig. 2 is a perspective view of an embodiment of the invention having four pipette channels. A syringe pump (not shown in Figs.) or other type of mechanism that can realize fluid volume exchange is connected to the inlet 101 e of the manifold 101 via tubing 108. The four outlets of the manifold 101 are coupled to four respective, disposable tip holders (one is labeled 101d in Fig. 2). The tip holders as shown in Fig. 2 have threaded connectors, and external unthreaded sleeves that firmly abut the housing of the manifold 101, and tightly fit into the disposable pipette tips forming seals to minimize air leakage. A disposable tip 102d is shown installed on tip holder lOld.

[0025] Double acting pneumatic cylinder 207 is fixed to the manifold 101 with a mounting plate 206. The cylinder shaft 204 is fixed with the disposable tip ejector 106 by nuts threaded onto cylinder shaft 204. When the cylinder shaft 204 is pushed out, the tip ejector 106 simultaneously pushes down the disposable tips 102 from their respective holders 101. Mounting plate 206 also supports an auxiliary locating rod 203 that prevents the remover 106 from rotating around the axis the cylinder shaft 204 when it moves up-and-down.

[0026] In the embodiment shown in cross-section in Fig. 3, the air inlet of the manifold diverter is connected with its air outlet through a buffer zone 101f. The buffer zone 10 If can be formed by a deep blind, transverse hole that is bored into the side of the manifold and then sealed with a plug 202 as shown. The radial or cross-sectional dimension of the buffer zone 101 f should be larger than that of each outlet orifice of the manifold, to avoid or reduce the performance differences between channels when they aspirate or dispense fluid due to differences in resistance to air flow, so a buffer effect is achieved. The buffer zone has the function of dispensing fluid equally, based on the volume exchange principle, it can equally dispense a volume of air aspirated and expelled by the syringe pump into each pipetting channel, resulting in consistency of pipetting operation of each channel. As discussed above, the manifold can adopt other mechanisms to equally dispense fluid in other embodiments of the invention.

[0027] The automatic pipettor provided by the present utility model can be fixed onto the move up-and-down slide of the kinematic axis of a robot. The robot can then position the automatic pipettor to realize the operations such as loading/unloading a disposable tip, and aspirating and dispensing fluid samples automatically. The pipetting method provided by embodiments of the present invention can be applied to plate replication, sample dispensing, and sample mixing.

[0028] Fig. 4 illustrates an operation of an embodiment of the invention, in which a robot positions the automatic pipetter 271 above a holder 310 for disposable pipette tips 102. Next the robot lowers the autopipettor, pressing the tip holders into the disposable tips, so that tip holder and disposable tip are coupled by press fit and deformation of the disposable tip. Then, as shown in Fig. 5, the robot moves to an array of fluid samples 401 to aspirate. The disposable tips are then dipped by the robot into the respectively positioned fluid samples for aspiration. Next the robot takes the automatic pipettor to a target position and dispenses the samples. Finally, the robot can move the autopipettor over a waste or recycling receptacle into which the disposable pipette tips are ejected, as shown in Fig. 6, thereby completing the operation to move fluid samples from their respective sources to their respective destinations.

[0029] Before aspirating a sample fluid, the autopipettor can pre-aspirate a volume of air to promote subsequent, thorough dispensing of a subsequently aspirated fluid sample. Dispensing the pre-aspirate along with the fluid sample can prevent some fluid sample from remaining inside the cavity of the disposable tip, and therefore it achieves high precision of pipetting operation. [0030] The containers to hold sample can be any kind of well plates, such as 96 well plate, or the containers, such as test tube, arranged in the holes of a plate spaced at a certain horizontal and vertical hole to hole pitches. In such cases the distance between pipetting channels is equal to one of said hole to hole pitches, or its integer multiple.

[0031] In another embodiment of a method of the invention, a volume of sample fluid aspirated in a single step, can be dispensed as smaller volumes (equal, or individually different volumes) to a number of successive destinations. During sample dispensing, after dispensing fluid to one target position, a droplet may remain hanging on the outside of the disposable tip, which can affect the precision of subsequent dispensing operations thereafter due to the hanging droplet. The dispensing precision can be increased by controlling the syringe pump to aspirate such droplets into pipetting channels first and then dispensing the fluid.

[0032] Fluid samples that need to be mixed can be deposited in the same container. The robot can then dip a disposable tip of the autopipettor into the collection of fluid samples, and the autopipettor can be cycled to aspirate and dispense substantially all of the aggregate volume of the fluid samples so as to mix the samples by fluid turbulence during the aspiration and dispensation. The operation can be repeated as many times as necessary for thorough mixing.

[0033] Fig. 7 is a block diagram of a controller for an embodiment of the invention. Embedded microcontroller 704 comprises, for example, an 80C552 8-bit microprocessor from Philips Semiconductor with associated memory (not shown). Embedded microcontroller 704 communicates with actuator drive circuits 706 and 708 via microcomputer I/O and expansion circuit 705. Comparable, suitable alternative embodiments are readily identified by one of ordinary skill in the embedded microcontroller arts. Actuator driver circuit 708 controls actuator 103b for the syringe pump 103. Actuator driver circuit 706 controls a pressure valve for actuating pneumatic cylinder 207. Embedded microcontroller 704 communicates with a system operator via a personal computer 701 through serial communication port 702 and RS232 unit 703. Software controlling the fluid dispensing apparatus can execute on embedded microcontroller 704, personal computer 701, or both. A system operator selects program parameters for the operation of the fluid dispensing apparatus via a user interface on personal computer 701. In some embodiments, embedded microcontroller 704 may have rudimentary user interface features such as pushbutton controls, and/or status indicator displays.

[0034] The operation of controller 110 can be coordinated with the operation of the robotic positioning device to synchronize the dipping and positioning of the pipette tips with associated aspiration, dispensing, and purging operations such as described in the methods, above.

[0035] Variations and extensions of the embodiments described are apparent to one of ordinary skill in the art. For example, the manifold may have different (i.e. other than co-linear) configurations of output ports for the pipette tips, for example a rectangular array. Also, embodiments of the invention can be used to precisely dispense volumes of sample fluid for applications other than microarrays for biological and/or chemical testing. Other applications, features, and advantages of this invention will be apparent to one of ordinary skill in the art who studies this invention disclosure. Therefore the scope of this invention is to be limited only by the following claims.

Claims

1. A multichannel pipettor, comprising:

a volumetrically apportioned pressure dividing manifold having an inlet port and a plurality of outlet ports;

a pump that is capable of displacing a volume of air, and is coupled to the inlet port of the volumetrically apportioned pressure dividing manifold; and

a pipette tip coupled to each of the plurality of outlet ports of the volumetrically apportioned pressure dividing manifold;

wherein the volumetrically apportioned pressure dividing manifold apportions a substantially equal volume displacement of air to each of the plurality of pipette tips.

2. The multichannel pipettor of claim 1 , further comprising an electromechanical actuator for the pump.

3. The multichannel pipettor of claim 1 , wherein the pump comprises a syringe pump.

4. The multichannel pipettor of claim 1, further comprising a pipette tip ejection mechanism.

5. The multichannel pipettor of claim 4, further comprising a pneumatic actuator for actuating the pipette tip ejection mechanism.

6. The multichannel pipettor of claim 1 in which at least one of the plurality of pipette tips is coupled by a press fit onto the volumetrically apportioned pressure dividing manifold by a pipette tip holder.

7. The multichannel pipettor of claim 6 in which the pipette tip holder is formed integrally with the volumetrically apportioned pressure dividing manifold.

8. The multichannel pipettor of claim 6 in which the pipette tip holder has a threaded section formed thereon, and is assembled with the volumetrically apportioned pressure dividing manifold with a threaded connection.

9. The multichannel pipettor of claim 1 wherein a pipette tip is integrally formed with the volumetrically apportioned pressure dividing manifold.

10. The multichannel pipettor of claim 1 in which the volumetrically apportioned pressure dividing manifold further comprises a cavity communicating the inlet port with the plurality of outlet ports, said cavity having a cross-sectional dimension larger than a cross-sectional dimension of one of the plurality of outlet ports.

11. The multichannel pipettor of claim 1 in which the volumetrically apportioned pressure dividing manifold is coupled to a robotic positioner.

12. The multichannel pipettor of claim 1 wherein the operation of the multichannel pipettor and the operation of the robotic positioner are commonly controlled by a processor.

13. A multichannel pipettor, comprising:

means for displacing a volume of air into an inlet port;

means for substantially equally dividing the displaced volume of air among a plurality of outlet ports; and

fluid sample containment means, coupled to each of said plurality of outlet ports.

14. The multichannel pipettor of claim 13, further comprising means to eject the fluid sample containment means, coupled to each of said plurality of outlet ports, from the respective one of said plurality of outlet ports to which it is coupled.

15. The multichannel pipettor of claim 13 , further comprising automatic control means.

16. A method for operating a multichannel pipettor to promote the complete dispensing of a fluid sample under automatic control, comprising:

aspirating a volume of air into a pipette tip of a multichannel pipettor responsive to a first actuation of a syringe pump, the pipette tip and the syringe pump being in pressure communication via a volumetrically apportioned pressure dividing manifold;

robotically positioning the pipette tip of the multichannel pipettor in a receptacle containing a fluid sample;

aspirating a volume of fluid sample into a pipette tip responsive to a second actuation of the syringe pump;

robotically positioning the pipette tip over a destination receptacle;

expelling the volume of fluid sample from the pipette tip, followed by the volume of air from the pipette tip responsive to a third actuation of the syringe pump.

17. A method for operating a multichannel pipettor to enhance the volumetric precision of successive dispensations of a fluid sample, comprising:

robotically positioning a pipette tip of the multichannel pipettor in a receptacle containing a fluid sample; aspirating a volume of fluid sample into the pipette tip responsive to a first actuation of a syringe pump, the pipette tip and the syringe pump being in pressure communication via a volumetrically apportioned pressure dividing manifold;

robotically positioning the pipette tip over a first destination receptacle;

expelling a first part of the volume of fluid sample from the pipette tip responsive to a second actuation of the syringe pump;

aspirating a residual droplet of the fluid sample from the outside of the pipette tip responsive to a third actuation of the syringe pump;

robotically positioning the pipette tip over a second destination receptacle; and

expelling a second part of the volume of fluid sample from the pipette tip responsive to a fourth actuation of the syringe pump.

18. A method for automatically operating a multichannel pipettor to mix volumes of fluid samples, comprising:

robotically positioning a pipette tip in a receptacle containing the volumes of the fluid samples to be mixed;

aspirating at least a first part of the volumes of the fluid samples to be mixed into the pipette tip, responsive to a first actuation of a syringe pump, the pipette tip and the syringe pump being in pressure communication via a volumetrically apportioned pressure dividing manifold; and

expelling at least a second part of the aspirated at least a first part of the volumes of the fluid samples back into the receptacle, responsive to a second actuation of a syringe pump; wherein the fluid turbulence of the aspirating and expelling substantially mixes the volumes of the fluid samples.