EP1412087A1 - Systeme de distribution d'echantillons liquides microfabrique a deux broches - Google Patents

Systeme de distribution d'echantillons liquides microfabrique a deux broches

Info

Publication number
EP1412087A1
EP1412087A1 EP02744565A EP02744565A EP1412087A1 EP 1412087 A1 EP1412087 A1 EP 1412087A1 EP 02744565 A EP02744565 A EP 02744565A EP 02744565 A EP02744565 A EP 02744565A EP 1412087 A1 EP1412087 A1 EP 1412087A1
Authority
EP
European Patent Office
Prior art keywords
liquid sample
tip
pin
dispensing system
sample dispensing
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP02744565A
Other languages
German (de)
English (en)
Other versions
EP1412087A4 (fr
Inventor
John Gilbert
John Harley
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Cytonome Inc
Original Assignee
Cytonome Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from US10/029,108 external-priority patent/US6808683B2/en
Priority claimed from US10/027,516 external-priority patent/US20020197733A1/en
Priority claimed from US10/027,922 external-priority patent/US20030077570A1/en
Priority claimed from US10/028,852 external-priority patent/US7179423B2/en
Priority claimed from US10/027,484 external-priority patent/US20030015425A1/en
Priority claimed from US10/027,171 external-priority patent/US7041257B2/en
Application filed by Cytonome Inc filed Critical Cytonome Inc
Publication of EP1412087A1 publication Critical patent/EP1412087A1/fr
Publication of EP1412087A4 publication Critical patent/EP1412087A4/fr
Withdrawn legal-status Critical Current

Links

Classifications

    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/02—Burettes; Pipettes
    • B01L3/0241—Drop counters; Drop formers
    • B01L3/0244—Drop counters; Drop formers using pins
    • B01L3/0248—Prongs, quill pen type dispenser
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01D—SEPARATION
    • B01D57/00—Separation, other than separation of solids, not fully covered by a single other group or subclass, e.g. B03C
    • B01D57/02—Separation, other than separation of solids, not fully covered by a single other group or subclass, e.g. B03C by electrophoresis
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/02—Burettes; Pipettes
    • B01L3/0241—Drop counters; Drop formers
    • B01L3/0244—Drop counters; Drop formers using pins
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J2219/00—Chemical, physical or physico-chemical processes in general; Their relevant apparatus
    • B01J2219/00274—Sequential or parallel reactions; Apparatus and devices for combinatorial chemistry or for making arrays; Chemical library technology
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/02—Adapting objects or devices to another
    • B01L2200/026—Fluid interfacing between devices or objects, e.g. connectors, inlet details
    • B01L2200/027—Fluid interfacing between devices or objects, e.g. connectors, inlet details for microfluidic devices
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/14—Process control and prevention of errors
    • B01L2200/143—Quality control, feedback systems
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/14—Process control and prevention of errors
    • B01L2200/143—Quality control, feedback systems
    • B01L2200/147—Employing temperature sensors
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/089—Virtual walls for guiding liquids
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/04—Moving fluids with specific forces or mechanical means
    • B01L2400/0403—Moving fluids with specific forces or mechanical means specific forces
    • B01L2400/0415—Moving fluids with specific forces or mechanical means specific forces electrical forces, e.g. electrokinetic
    • B01L2400/0421—Moving fluids with specific forces or mechanical means specific forces electrical forces, e.g. electrokinetic electrophoretic flow
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/04—Moving fluids with specific forces or mechanical means
    • B01L2400/0403—Moving fluids with specific forces or mechanical means specific forces
    • B01L2400/0433—Moving fluids with specific forces or mechanical means specific forces vibrational forces
    • B01L2400/0439—Moving fluids with specific forces or mechanical means specific forces vibrational forces ultrasonic vibrations, vibrating piezo elements
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
    • G01N35/10—Devices for transferring samples or any liquids to, in, or from, the analysis apparatus, e.g. suction devices, injection devices
    • G01N2035/1027—General features of the devices
    • G01N2035/1034—Transferring microquantities of liquid
    • G01N2035/1037—Using surface tension, e.g. pins or wires
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
    • G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
    • G01N27/416—Systems
    • G01N27/447—Systems using electrophoresis

Definitions

  • the present invention relates to a liquid dispensing system for forming and dispensing droplets of a liquid sample.
  • sample handling system such as a microfluidic system capable of handling and analyzing chemical and biological specimens, quickly, efficiently and in a highly controllable manner.
  • a liquid sample to a microfluidic system is accomplished through sample channels or sample wells.
  • a capillary tube may be provided, which dispenses a liquid sample to a sample well, sample channel or other sample introduction port.
  • a - significant drawback of using a capillary tube concerns the low injection efficiency inherent to capillary tubes, that is, the ratio between the volume of liquid required for a particular chemical operation in a part of the microfluidic system, and the total volume of liquid required for the introductory operation. Moreover, it is generally difficult to control the precise volume of dispensed sample using capillary tubes. Furthermore, capillary tubes are subject to contamination, because the same port used to fill the tube is also used to eject the liquid sample.
  • U.S. Patent Number 6,101,946 of Martinsky describes a pin-based system for printing microarrays of biochemical substances.
  • the microarray printing system comprises a stainless steel printing pin having a sample channel and a flat tip that is machined with an electronic discharge machine (EDM).
  • EDM electronic discharge machine
  • the pin applies a biochemical substance by filling the sample channel and subsequently directly contacting a printing substrate, to deliver the sample from the sample channel to the printing substrate.
  • a drawback of the pin-based system described in the '946 patent concerns the ability to control the amount of delivered sample.
  • the pin-based system is subject to contamination and breakage, because it requires direct contact between the pin tip and the printing substrate.
  • Another drawback concerns the difficulty of precisely positioning the tip of the pin to provide sufficient contact to result in delivery of a sample.
  • U.S. Patent Number 6,110,426 of Shalon et al. describes a capillary dispenser for forming microarrays of biological samples.
  • the capillary dispenser comprises an elongate open capillary channel adapted to hold a liquid sample.
  • the channel is formed by a pair of spaced- apart, coextensive, elongate members, which are tapered toward one another and converge at a tip region at the lower end of the channel.
  • the elongate members are fixed relative to each other and the capillary channel is limited to a fixed volume. Furthermore, it is difficult to control the amount of sample that is acquired and dispensed from the capillary dispenser of the '246 patent.
  • the present invention provides a sample dispensing system comprising two microfabricated interacting pins for forming and dispensing droplets of a liquid sample.
  • Each pin includes a tip spaced predetermined distance from the other pin to define a sample acquisition region.
  • the pins acquire and hold a droplet of the liquid sample in the sample acquisition region formed in the space between the tips and apply the droplet to a selected sample handing system.
  • the distance between the tips is variable to accommodate different liquid samples having varying physical properties and to vary the volume of the acquired droplet.
  • a droplet dispensing system comprises two separately movable pins for holding a droplet of a liquid sample.
  • the droplet dispensing system comprises a holder, a first pin connected to the holder and having a first tip and a second pin connected to the holder and having a second tip spaced an initial separation distance from the first tip to form a sample acquisition region for holding a predetermined volume of liquid sample.
  • the initial separation distance is variable.
  • a method of applying a droplet of liquid sample to a substrate comprises providing a dispensing system comprising two pins separated by a variable distance, immersing the pin tips in a reservoir to acquire a droplet of a liquid sample, and contacting the substrate to deposit a spot of the liquid sample on the substrate.
  • the spot has a predetermined volume less than the volume of the droplet.
  • a method of diluting a first liquid sample in a second liquid comprises providing a dispensing system comprising two pins having a first tip and a second tip separated from the first tip by a variable distance, acquiring a droplet of the first liquid sample between the pins and immersing the pin tips containing the droplet of the first liquid sample in a second reservoir containing a second liquid, whereby the droplet of the first liquid sample is diffused into the second liquid.
  • a two-pin droplet dispensing system comprises a holder, a first pin connected to the holder and having a first tip and a second pin connected to the holder and having a movable tip spaced a predetermined distance from the first tip to form a sample acquisition region for holding a predetermined volume of liquid sample.
  • the system further comprises a driver for effecting movement of the movable tip with respect to the first tip.
  • a liquid sample dispensing system comprising a holder a first pin having a first tip and a relaxation region for absorbing an impact on the tip is provided.
  • Figures la and lb illustrate the two-pin dispensing system of an illustrative embodiment in a sample acquisition mode.
  • Figure 2 illustrates the two-pin dispensing system of an illustrative embodiment in a spotting mode.
  • Figures 3 a and 3b illustrate the two-pin dispensing system of an illustrative embodiment in a dilution mode.
  • Figure 4 illustrates an alternative embodiment of the two-pin dispensing system, including a fixed pin and a movable pin.
  • Figure 5 is a detailed view of the fulcrum region of the two-pin dispensing system of Figure 4.
  • Figure 6 is a detailed view of the driver of the two-pin dispensing system of Figure 4.
  • Figures 7a, 7b and 7c are detailed views of the tip region of the two pins of the pin dispensing system of Figure 4.
  • Figure 8 illustrates the relaxation region of the pin dispensing system of Figure 4.
  • Figure 9 is a scanning electron microscope (SEM) image of an array of two-pin dispensing systems that are microfabricated from a silicon wafer according to the teachings of the illustrative embodiment of the present invention.
  • Figure 10 is a SEM image showing a detailed view of a tip region of one of the two-pin dispensing systems of Figure 9.
  • Figure 11 is a SEM image illustrating an array of two-pin dispensing systems having relaxation regions that are microfabricated from a silicon wafer according to the teachings of the illustrative embodiment of the present invention.
  • the present invention provides a dispensing system for dispensing a predetermined volume of liquid sample.
  • the dispensing system of the present invention provides precise acquisition and delivery of a liquid sample to a sample handling system using two interacting pins.
  • the dispensing system is suitable for use in a basic research or a commercial environment.
  • the dispensing system significantly improves sample introduction into a sample handling system by increasing the efficiency, speed and controllability of forming and dispensing droplets while significantly reducing waste and contamination.
  • the invention will be described below relative to an illustrative embodiment. Those skilled in the art will appreciate that the present invention may be implemented in a number of different applications and embodiments and is not specifically limited in its application to the particular embodiment depicted herein.
  • Figures la and lb illustrate a two-pin dispensing system 10 of an illustrative embodiment of the present invention in a sample acquisition mode for acquiring a droplet of a liquid sample having a predetermined volume from a reservoir containing a supply of the liquid sample.
  • the illustrative two-pin dispensing system 10 comprises a pair of separately movable, interacting pins sized and configured to hold a droplet of liquid between the tips of the pins.
  • the two-pin dispensing system 10 comprises a first pin 11 and a second pin 12, which are movably connected to a holder 13.
  • the tips 11a, 12a are separated by an initial separation distance D to form a sample acquisition region
  • sensors 16 are provided on one or more of the pins 11, 12 to measure the separation distance D between the pin tips 11a, 12a.
  • the holder of the invention can include any suitable structure for retaining or holding the pins.
  • the pin tips 1 la, 12a are immersed in a reservoir 17 containing a supply of a selected liquid sample.
  • the pin tips 11a, 12a are positioned to allow capillary flow into to sample acquisition region 14.
  • the capillary force induced in the sample acquisition region 14 pulls a droplet 18, having a volume defined by the separation distance of the pin tips 11a, l ib, into the sample acquisition region 14.
  • the capillary force produced between the surfaces of the pin tips holds the droplet in the sample acquisition region 14 formed between the two pin tips 11a, l ib.
  • the actuators 15 in the pins 11, 12 move the pins to vary the separation distance D between the tips, thereby varying the amount of sample that is acquired by the two-pin dispensing system, or to dispense the sample therefrom.
  • the two-pin dispensing system 10 is configured to acquire liquid samples in volumes between about fifty picoliters and about fifty nanoliters.
  • the acquired volume is not limited to this range and that the pins may be spaced apart to accommodate any suitable volume of liquid.
  • the actuators 15 can also compensate for varying physical properties of the particular liquid sample, such as viscosity, surface tension, and the like, by modifying the separation distance D between the pins.
  • the sensors 16 may also be utilized to measure the force applied between the tips and the physical properties of the acquired liquid sample on the fly. In this manner, the settings (i.e. the pin separation distance) of the pin dispensing system 10 can be modified to compensate for variations in the measured properties of the liquid sample in real time.
  • the droplet dispensing system 10 is fabricated from a silicon wafer using a microfabrication technique, such as a standard photolithography etching process, to fabricate the pin structures.
  • a microfabrication technique such as a standard photolithography etching process
  • the pin dispensing system may be made out of glass, plastic or any other suitable material.
  • an array of droplet dispensing systems 10, each comprising two pins having a variable separation distance may be formed on a single substrate, such as a silicon wafer.
  • an array of up to about 300 or more two-pin dispensing systems 10 may be formed on a four-inch silicon wafer.
  • Figure 2 illustrates the two-pin dispensing system 10 of the illustrative embodiment in a spotting mode.
  • the two-pin dispensing system 10 may be utilized as a spotting system for printing or discharging arrays of biochemicals, such as nucleic acid molecules or proteins, or other suitable liquid samples to a sample handing system, such as a printing substrate, titre plate, microfluidic system or device, and the like for use in proteomics, genomics, screening, diagnostics and other applications.
  • a sample handing system such as a printing substrate, titre plate, microfluidic system or device, and the like for use in proteomics, genomics, screening, diagnostics and other applications.
  • the dispensing system After the dispensing system acquires a droplet, the dispensing system is moved in close proximity to a surface 20.
  • the surface 20 may comprise a solid surface or a liquid.
  • the surface 20 may comprise a porous structure, such as a porous membrane, or a non-porous structure, such as a microscope slide.
  • the loaded pins deposit a spot 21 on the surface 20 having a selected spot volume by direct contact between the pin tips 11a, 1 lb and the surface.
  • the separation distance D2 during contact may be varied to increase or decrease the volume of the dispensed spot of the liquid sample.
  • the volume of the dispensed spot 21 is significantly smaller than the volume of the acquired droplet 18, and is generally sub-nanoliter in volume, though one skilled in the art will recognize that the invention is not limited to this range.
  • the use of the two-pin dispensing system of the illustrative embodiment in spotting applications provided enhanced control over the size of the deposited spots in a microarray, and also allows for smaller spots to be formed and deposited.
  • the pin dispensing system may further be utilized as a wet deposit system with dilution to dilute a selected volume of a first liquid in a second liquid sample.
  • Figures 3 a and 3b illustrate the two-pin dispensing system 10 in a dilution mode, wherein the acquired droplet 18 of a sample is diluted in a larger supply of a target fluid 30. After the dispensing system 10 acquires a droplet 18, the size of which is defined by the separation distance of the pin tips 11a, 12a, the pin tips 11a, 12a are immersed in a reservoir 30 containing a target fluid. The droplet 18 automatically dilutes into the target fluid via mixing and diffusion.
  • Figure 4 illustrates a two-pin dispensing system 40 having a fixed pin and a movable pin according to an alternate embodiment of the invention.
  • the resting position of a first pin 41 is fixed relative to a substrate 43 and the resting position of the second pin 42 is movable relative to the first pin 41 and the substrate 43.
  • the two-pin dispensing system 40 further includes a driver 44 for varying the separation distance between the tips 41a, 42a by adjusting the position of the second movable pin 42 in a fulcrum region 46.
  • the movable pin rotates about a fixed pivot point 45 under the control of the driver 44 to adjust the separation distance at the tips.
  • the pins 41 , 42 further include a relaxation region 51 for preventing breakage of the tips.
  • the relaxation region 51 may be formed in one or both of the pins 41, 42 of the two-pin dispensing system 40.
  • the illustrative two-pin dispensing system 40 is formed from a silicon wafer using a standard photolithography etching process to fabricate the pins 41, 42, the relaxation region 51 , the driver 44, and the fixed point 45 of the fulcrum region 46 in the wafer substrate 43.
  • the two-pin dispensing system 40 is fabricated from a silicon wafer having dimensions of about one square centimeter.
  • the pins 41,42 have a length of about five millimeters, though one skilled in the art will recognize that the invention is not limited to this size.
  • a larger silicon wafer or other suitable substrate is provided, and an array of pin dispensing systems is fabricated on the larger silicon wafer.
  • a silicon wafer having a size of about ten square centimeters may be used to fabricate an array of about seventy two-pin dispensing systems 40 thereon.
  • a fifteen square centimeter silicon wafer can be utilized to fabricate over one hundred two-pin dispensing systems 40 in the silicon wafer substrate.
  • any suitable configuration can be employed to move one or both of the pins.
  • FIG. 5 is a detailed view of the fulcrum region 46 of the two-pin dispensing system of Figure 4.
  • the movable pin 42 is configured to pivot about a fixed point 45 to vary the separation distance of the two pin tips.
  • the driver 44 applies a force to an application region 47 of the fulcrum region 46 to cause the movable pin 42 to rotate, thereby effecting movement of the movable pin tip 42a relative to the tip 41a of the fixed pin 41.
  • the fulcrum region 46 includes gaps 48 are formed in the substrate 43 adjacent to the fixed point 45 to allow for rotation of the pin 42 about the fixed point in response to activation of the driver 44.
  • the fulcrum region further includes bending sensors, illustrated as piezoresistors 62, on the left and right side of the fulcrum region to allow differential sensing of actual bending of the pin 42 in the fulcrum region.
  • bending sensors illustrated as piezoresistors 62
  • the amount of bending, and the resultant tip separation distance may be controlled using a closed loop feedback system.
  • the use of bending sensors further limits nonlinear temperature effects by allowing real-time sensing of tip displacement.
  • FIG 6 is a detailed view of the driver 44 of the two-pin dispensing system 40 of Figure 4.
  • the driver 44 comprises a bar of silicon that imparts a force on the application region 47 of the fulcrum 46 to move of tip 42a a predetermined amount.
  • the driver 44 expands a predetermined amount in response to a temperature increase.
  • the expansion of the driver 44 forces rotation of the fulcrum about the pivot point.
  • the system is configured such that the ratio between the amount of movement of the tip 42a in response to expansion of the driver 44 to the amount of expansion of the driver is greater than one hundred. In other words, a driver expansion of one micron causes a one hundred micron displacement of the pin tip 42a.
  • the driver 44 has an initial length L of four millimeters. A thirty-degree rise in temperature of the silicon results in a 1.08 micrometer expansion of the driver 44. The expansion of the driver 44 forces the pin 42 to rotate about the fixed pivot point 45, thereby increasing the separation distance between the tips 41a, 42a by greater than 108 microns.
  • heating resistors 49 are affixed to the driver for applying heat to the driver 44.
  • the heating resistors may comprise poly resistors, diffused resistors or any suitable means for applying heat to the driver 44 in order to effect controlled expansion of the driver 44 and to vary the separation distance between the tips 41a, 42a.
  • cooling fins are provided in the driver 44 near the fulcrum region 45 to prevent unwanted heating of the driver in the fulcrum region.
  • a temperature sensor in communication with the heating means is included in the two-pin dispensing system 40 to provide closed loop control of the driver 44 temperature.
  • the two-pin dispensing system is not limited to the illustrative driver.
  • the driver 44 comprises an electrostatic system, a piezoelectric system, an electromechanical system, a thermoelectric actuator or any suitable system for applying a predetermined and defined force to cause controlled adjustment of the separation distance between the pin tips 41a, 42a.
  • the two-pin dispensing system is not limited to a fulcrum for varying the separation distance and that any suitable mechanism for varying the separation distance may be utilized.
  • FIGs 7a and 7b are detailed views of the tip region of the two pins of the two- pin dispensing system 40 of Figure 4.
  • the tips 41a, 42a are spaced apart a predetermined distance D, which is defined by the driver 44.
  • Each tip includes a sample contact surface 70, defined by the tip height H and the tip depth S, which are fixed values determined by the shape of the sample surfaces 70.
  • the volume of the sample acquisition region 14 and thus the volume of an acquired sample droplet corresponds to the volume of the space defined between the tips, or the product of the tip height H, the tip depth S and the separation distance D.
  • the separation distance D between the pin tips 41a, 42a is between about twenty-five microns and about one hundred twenty five microns.
  • the resulting volume of a captured droplet is between about 250 picoliters and about 1.25 nanoliters.
  • a tip separation distance between about 25 and 125 microns results in an acquired droplet having a volume of between about 1.0 nanoliters and about 5 nanoliters. Tips having a depth of 500 microns and a height of 500 microns, form a droplet having a volume between about 6.25 nanoliters and about 31.5 nanoliters when the separation distance between the tips is between about 25 and about 125 microns.
  • the tip contact surfaces 70 defined by the tip heights H and tip depths S may form parallel faces or, according to a preferred embodiment, may be tapered, so that the separation distance D is reduced towards the bottom and/or front of the tip surface. In this manner, smaller droplet volumes may be accommodated.
  • the slope of the tips 41a, 42a may be varied in regions 71 and 72 to improve droplet shape and enhance delivery of the droplet.
  • Figure 7c is a cross-sectional view of the tips 41a, 42a according to an alternate embodiment.
  • the tip surfaces 70' are curved to hold form a cylindrical or conical sample acquisition region 14 therebetween.
  • the one or both of the tip surfaces 70 and/or the outside shaft surface are coated with a hydrophilic, hydrophobic or other chemical coating to enhance droplet acquisition and dispensing.
  • the tips 41, 42 may be formed of or coated with a hydrophilic coating to enhance retention of a sample in the sample acquisition region.
  • the outside shaft surfaces of the tips 41,42 are coated with gold or another suitable hydrophobic material without affecting the tip surfaces 70 defining the sample acquisition region 14.
  • the use of a metal coating provides enhanced control over the volume and release of a droplet.
  • the use of silicon and/or gold additionally allows for more vigorous cleaning solutions to be utilized when cleaning the tips without degrading the system. In this manner, contamination of the tips is reduced.
  • the coating may be applied in a pattern to the tip surfaces 70 or the other surfaces of the tips 41, 42 by shadow masking.
  • the coating may be sputtered, or evaporated on a surface in a predetermined pattern, defined by a mask.
  • any suitable pattern for directing the liquid sample and enhancing control over sample acquisition and dispensing may be utilized.
  • the dispensing system may comprise a single pin having a suitable pattern coating applied to the surfaces of the pin tip.
  • the shaft of the single pin may be coated with a suitable hydrophobic coating and the tip of the pin may be coated with a suitable hydrophilic coating to enhance acquisition and dispensing of a liquid sample.
  • Figure 8 illustrates the relaxation region 51 of the pin dispensing system 40 of Figure 4.
  • the pin tips 41, 42 are brittle and subject to breakage when accidentally touched down to surfaces, due to their size and the material used to fabricate the pins.
  • the illustrative relaxation region 51 comprises a spring 52 formed between the tip 41a and the substrate 43. When the tip 41a contacts a surface, the spring absorbs the impulse and retracts the tip 41a to prevent breakage.
  • the springs 52 in the pins are configured to move the corresponding tip up and away from the other tip to prevent collision of the tips.
  • the invention is not limited to the illustrative spring design. One skilled in the art will recognize that any suitable spring design may be utilized to form the relaxation region 51 to protect the pin tips from breakage.
  • the spring 52 includes sensors to measure of the force of contact between the tip and a surface.
  • sensors to measure of the force of contact between the tip and a surface.
  • differential piezoresistive sensors may be included in the spring 52 and connected to an actuator (not shown) to control the spring using feedback control loop.
  • the spring sensor may also be utilized to measure the force exerted by the droplet on the tips, and allow the driver to compensate for variable forces exerted by the droplet on the tips.
  • a relaxation region may be implemented in a two-pin dispensing system comprising a pair of spaced- apart, fixed pins defining a sample acquisition region of fixed volume.
  • the two-pin dispensing system 10 or 40 of the illustrative embodiment may be microfabricated from a suitable substrate, such as silicon, glass or plastic.
  • a suitable substrate such as silicon, glass or plastic.
  • photolithography may be utilized to form the pin structures in the substrate.
  • the pattern of the two pins and other components of the two-pin dispensing system 10 or 40 are imprinted on a silicon wafer, or other substrate, using one or more photoresist layers that are patterned by UV or other light projected through one or more photo-masks containing the pattern on it.
  • the substrate is then etched to fabricate the two-pin structure.
  • any suitable microfabrication technique may be utilized to manufacture the two-pin dispensing system of the illustrative embodiment of the present invention.
  • microfabrication technique may further be utilized to fabricate single-pin dispensing systems from a silicon wafer or other suitable substrate.
  • Figure 9 is a scanning electron microscope (SEM) image of an array of two-pin sample dispensing systems 100 according to an embodiment of the invention and formed from a silicon wafer 101 using the above-described microfabrication technique. As shown, a plurality of two-pin dispensing systems are fabricated from a single silicon wafer substrate. Each two-pin dispensing system comprises a pair of elongated pins that are spaced apart to define a sample acquisition region between the tips of the pins.
  • Figure 10 is another SEM image showing a detailed view of the tip region of one of the microfabricated two-pin dispensing systems of Figure 9. As shown, the pins are etched in a silicon wafer to define a sample acquisition region 140 between the tips of the pins. As illustrated, the microfabricated pin tips have a separation distance of less than about 100 microns.
  • Figure 11 is another SEM image of an array of microfabricated two-pin sample dispensing systems 110 having relaxation regions 51 according to an embodiment of the invention.
  • the array is also formed from a silicon wafer 101 using the above- described microfabrication technique.
  • the relaxation region 51 is formed by etching the silicon wafer in the region between the pin tips and the holder to define a spring for absorbing an impact on the tips.
  • the relaxation region 51 prevents breakage of the pin tips 41, 42 when the pin tips contact a surface.
  • the two-pin dispensing system provides significant improvements to the process of forming and dispensing droplets of samples for spotting and dilution applications.
  • the illustrative configuration provides precise control over the amount of liquid sample that is acquired and deposited through the use of two pins having a variable separation distance. Adjusting the separation distance between the pin tips easily and precisely modifies the volume of the acquired liquid droplet and the deposited liquid droplet. Furthermore, measurements of the physical properties of the liquid volume can be made on the fly and the tip separation can be modified quickly and easily to compensate for variations.
  • the use of sensors provides precise control of the tip separation distance to optimize the process of acquiring and dispensing droplets of a liquid sample.

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  • Chemical & Material Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Engineering & Computer Science (AREA)
  • Organic Chemistry (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Molecular Biology (AREA)
  • Clinical Laboratory Science (AREA)
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  • Bioinformatics & Cheminformatics (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Sampling And Sample Adjustment (AREA)
  • Automatic Analysis And Handling Materials Therefor (AREA)
  • Feeding, Discharge, Calcimining, Fusing, And Gas-Generation Devices (AREA)

Abstract

L'invention concerne un système de distribution d'échantillons liquides à deux broches (10) qui comprend une paire de broches mobiles séparément (11, 12) destinées à recevoir une gouttelette de liquides entre elles. Chaque broche est pourvue d'une pointe (11a, 12a) espacée d'une distance prédéterminée par rapport à l'autre broche afin de définir une zone de prélèvement d'échantillons (14). Ces broches prélèvent et retiennent une gouttelette de l'échantillon liquide dans la zone de prélèvement d'échantillons formée dans l'espace situé entre les pointes et appliquent la gouttelette sur un système choisi de manipulation d'échantillons. La distance qui sépare les pointes peut varier afin de recevoir différents échantillons liquides présentant des propriétés physiques variables et afin de modifier le volume de la gouttelette prélevée.
EP02744565A 2001-06-20 2002-06-20 Systeme de distribution d'echantillons liquides microfabrique a deux broches Withdrawn EP1412087A4 (fr)

Applications Claiming Priority (17)

Application Number Priority Date Filing Date Title
US27484 1993-03-03
US29951501P 2001-06-20 2001-06-20
US299515P 2001-06-20
US32500101P 2001-09-25 2001-09-25
US325001P 2001-09-25
US10/029,108 US6808683B2 (en) 2001-09-25 2001-12-21 Droplet dispensing system
US10/027,516 US20020197733A1 (en) 2001-06-20 2001-12-21 Microfluidic system including a virtual wall fluid interface port for interfacing fluids with the microfluidic system
US10/027,922 US20030077570A1 (en) 2001-09-20 2001-12-21 Small molecule substrate based enzyme activity assays
US27922 2001-12-21
US10/028,852 US7179423B2 (en) 2001-06-20 2001-12-21 Microfluidic system including a virtual wall fluid interface port for interfacing fluids with the microfluidic system
US27171 2001-12-21
US27516 2001-12-21
US29108 2001-12-21
US10/027,484 US20030015425A1 (en) 2001-06-20 2001-12-21 Microfluidic system including a virtual wall fluid interface port for interfacing fluids with the microfluidic system
US28852 2001-12-21
US10/027,171 US7041257B2 (en) 2001-09-25 2001-12-21 Microfabricated two-pin liquid sample dispensing system
PCT/US2002/019924 WO2003000422A1 (fr) 2001-06-20 2002-06-20 Systeme de distribution d'echantillons liquides microfabrique a deux broches

Publications (2)

Publication Number Publication Date
EP1412087A1 true EP1412087A1 (fr) 2004-04-28
EP1412087A4 EP1412087A4 (fr) 2005-03-09

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EP02744565A Withdrawn EP1412087A4 (fr) 2001-06-20 2002-06-20 Systeme de distribution d'echantillons liquides microfabrique a deux broches

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EP (1) EP1412087A4 (fr)
JP (1) JP2005502448A (fr)
CA (1) CA2451201A1 (fr)
WO (1) WO2003000422A1 (fr)

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US7153699B2 (en) * 2001-12-21 2006-12-26 Cytonome, Inc. Microfabricated two-pin system for biomolecule crystallization
US7258839B2 (en) 2001-12-21 2007-08-21 Cytonome, Inc. Temperature controlled microfabricated two-pin liquid sample dispensing system
JP4624133B2 (ja) * 2004-03-29 2011-02-02 京セラ株式会社 液体印刷ピン
KR20150022751A (ko) * 2012-03-16 2015-03-04 라이프 테크놀로지스 코포레이션 생물학적 반응 시스템을 위한 코팅된 기판
EP3502650B1 (fr) * 2017-12-22 2023-07-19 Universiteit Maastricht Procédé et appareil pour appliquer un liquide sur un substrat
JP7475953B2 (ja) 2020-05-01 2024-04-30 キヤノンメディカルシステムズ株式会社 自動分析装置

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See also references of WO03000422A1 *

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EP1412087A4 (fr) 2005-03-09
JP2005502448A (ja) 2005-01-27
WO2003000422A1 (fr) 2003-01-03
CA2451201A1 (fr) 2003-01-03

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