EP3259064B1 - Double séringue pour amener un liquide dans un système micro-fluidique - Google Patents

Double séringue pour amener un liquide dans un système micro-fluidique Download PDF

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Publication number
EP3259064B1
EP3259064B1 EP16708075.3A EP16708075A EP3259064B1 EP 3259064 B1 EP3259064 B1 EP 3259064B1 EP 16708075 A EP16708075 A EP 16708075A EP 3259064 B1 EP3259064 B1 EP 3259064B1
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Prior art keywords
chamber
outer chamber
holes
piston
fluid
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German (de)
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EP3259064A1 (fr
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Gerhard Hartwich
Norbert Persike
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Friz Biochem Gesellschaft fuer Bioanalytik mbH
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Friz Biochem Gesellschaft fuer Bioanalytik mbH
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    • 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
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
    • B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip
    • B01L3/502715—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces
    • 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
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
    • B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip
    • B01L3/50273—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip characterised by the means or forces applied to move the fluids
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L7/00—Heating or cooling apparatus; Heat insulating devices
    • B01L7/52—Heating or cooling apparatus; Heat insulating devices with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples
    • 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
    • B01L2200/02—Adapting objects or devices to another
    • B01L2200/028—Modular arrangements
    • 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
    • B01L2200/16—Reagents, handling or storing thereof
    • 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
    • B01L2300/06—Auxiliary integrated devices, integrated components
    • B01L2300/0627—Sensor or part of a sensor is integrated
    • B01L2300/0636—Integrated biosensor, microarrays
    • 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
    • B01L2300/06—Auxiliary integrated devices, integrated components
    • B01L2300/0627—Sensor or part of a sensor is integrated
    • B01L2300/0645—Electrodes
    • 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
    • B01L2300/0809—Geometry, shape and general structure rectangular shaped
    • B01L2300/0816—Cards, e.g. flat sample carriers usually with flow in two horizontal directions
    • 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
    • B01L2300/0861—Configuration of multiple channels and/or chambers in a single devices
    • B01L2300/087—Multiple sequential chambers
    • 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
    • B01L2400/0475—Moving fluids with specific forces or mechanical means specific mechanical means and fluid pressure
    • B01L2400/0478—Moving fluids with specific forces or mechanical means specific mechanical means and fluid pressure pistons
    • 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
    • B01L2400/06—Valves, specific forms thereof
    • B01L2400/0677—Valves, specific forms thereof phase change valves; Meltable, freezing, dissolvable plugs; Destructible barriers
    • 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
    • B01L3/021—Pipettes, i.e. with only one conduit for withdrawing and redistributing liquids
    • B01L3/0217—Pipettes, i.e. with only one conduit for withdrawing and redistributing liquids of the plunger pump type
    • B01L3/0231—Pipettes, i.e. with only one conduit for withdrawing and redistributing liquids of the plunger pump type having several coaxial pistons
    • 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
    • B01L3/52—Containers specially adapted for storing or dispensing a reagent
    • B01L3/527—Containers specially adapted for storing or dispensing a reagent for a plurality of reagents

Definitions

  • the invention relates to a double syringe for the supply of a liquid in a microfluidic system and a microfluidic cartridge with such a double syringe.
  • nucleic acid also encompasses nucleic acid sequences in the context of this description.
  • Powerful and rapid detection methods employ an array technology using so-called DNA chips, which enable a surface-sensitive detection of nucleic acid oligomer hybridization events.
  • DNA chips which enable a surface-sensitive detection of nucleic acid oligomer hybridization events.
  • the actual, anyway highly sensitive detection is preceded by an amplification step in which the nucleic acids to be detected are amplified so that they are ultimately present in a concentration above the detection limit of the selected detection method.
  • PCR polymerase chain reaction
  • the PCR is an enzymatic reaction for the amplification of nucleic acids, which are essentially aqueous or liquid reaction mixture runs with very small volumes.
  • the reaction mixture contains a nucleic acid-containing sample and the primers, nucleotides and a polymerase which are also necessary for the reaction.
  • buffers and preferably divalent ions the reaction mixture is adjusted so that the optimum reaction conditions for the particular application prevail.
  • the PCR is based on a multi-repeating cycle of three steps of denaturation, hybridization, and extension that occur at different temperatures. In each cycle, the number of nucleic acids to be amplified is roughly doubled so that significant amplification can be achieved with a small number of cycles.
  • PCR in general, on Real Time PCR, in which the amount of nucleic acid is determined while the PCR reaction is underway, and on preferred methods for the electrochemical detection of nucleic acid oligomer hybridization events on test sites of a DNA chip is referred to pamphlet DE 10 2011 056 606 B3 the applicant, the disclosure of which is included in the present application in this respect.
  • Microfluidic devices for processing bioparticles are known, for example, from the publications EP1641564B1 and DE 10 2010 043 030 A1 known.
  • liquids are often supplied to a microfluidic device by means of a pump via a closable valve.
  • a pump via a closable valve.
  • the object of the invention is to provide a delivery device for the supply of a liquid into a microfluidic system, which overcomes the disadvantages of the prior art, and in particular allows a simple, inexpensive, reliable supply of a closed volume of liquid in a microfluidic system.
  • the feed device is also referred to as a double syringe in the context of this description.
  • a particularly simple construction is achieved if the two cylinder chambers are arranged concentrically with a common cylinder axis.
  • the inner piston of the double syringe on its proximal side on a handle member which is gripped by an associated piston rod for axial movement of the inner piston.
  • the piston rod may in particular represent an external element which is brought together with the use of the double syringe with this.
  • the inner piston can also have on its proximal side a cavity into which the distal end of an associated piston rod can engage for the axial movement of the inner piston.
  • the inner piston of the double syringe is preferably provided with a rubber seal for closing the through-holes.
  • the closure position of the inner piston lies at the distal end of the cylindrical inner chamber, wherein the inner piston in the closed position bears substantially against the distal cylinder cover surface of the inner chamber.
  • the annular piston of the outer chamber is advantageously formed by a rubber ring.
  • the closure position of the annular piston is located at the distal end of the cylindrical outer chamber, the annular piston in the closed position abutting substantially on the distal cylinder top surface of the outer chamber.
  • the through-holes are expediently formed in the common cylinder wall of a cylindrical inner chamber and a cylindrical outer chamber, preferably in the distal end region of the cylinder wall at the same axial height circumferentially around the cylinder wall.
  • the storage space of the double syringe is already prefilled with a process liquid. Since the storage space is closed by the annular piston of the outer chamber, the process liquid can be provided in the double syringe storage stable.
  • a storage space for receiving chemicals, which is in particular prefilled with desired chemicals and a release function, such as a fusible wax layer or a magnetic particle cover layer.
  • the release function fixes and protects the chemicals in the storage room until they are released, for example by raising the temperature or by applying a suitable magnetic field.
  • This multi-fluid dual syringe may be further configured as described above with respect to the dual syringe for only one fluid.
  • the inner piston and the annular piston may advantageously be formed in the manner described above and each group Through hole may be formed at the same axial height circumferentially in the common cylinder wall of the two cylinder chambers.
  • the invention also includes a microfluidic cartridge for the detection of biomolecules in a sample solution having a microchannel system and a double syringe of the type described for supplying a process fluid into the microchannel system.
  • FIG. 1 illustrates the principle of detection of certain nucleic acids in a sample 92 present in a sample system 90 by a detection system 10.
  • the sample 92 may be, for example, a swab sample of a patient present at the end of a swab in a sample tube 90 and microbiologically MRSA strains (methicillin resistant Staphylococcus aureus) should be studied.
  • the detection system 10 includes a pumping device 20 having a process fluid 22 which is supplied to the sample 92 through a microchannel system 30 and an open valve 80 in a delivery step (flow direction 32).
  • the process fluid 22 passes through a storage area 70, in which it contains chemicals 72, in the exemplary embodiment, for example, for exposing the DNA of the sample 92, receives.
  • the detection chamber 40, the reaction chamber 50 and the pressure chamber 60 of the detection system 10 do not play any role in this feeding step.
  • the sample solution resulting from the lysis of the sample 92 in the sample system 90 is returned by the pumping device 20 to the detection system 10 (flow direction 34).
  • the sample solution is usually withdrawn into the pumping device 20 to ensure that at least the reaction chamber 50 of the detection system 10 is filled with sample solution.
  • the valve 80 is closed to the external sample system 90 to have a defined volume of fluid in the detection system 10 and to obtain a sealed system.
  • PCR polymerase chain reaction
  • the chemicals 52 required for this purpose are initially pre-assembled under a wax layer.
  • the wax layer is melted by an increase in temperature and thereby the PCR chemicals 52 are added to the sample solution in the reaction chamber 50.
  • the molten wax remains liquid or is taken up by multiple reservoirs in the periphery of the reaction chamber 50 to ensure that no solidified wax clogs adjacent microchannels 30.
  • the cyclic temperature profile in the reaction chamber 50 required for the PCR is generated by means of a heating / cooling device (not shown) associated with the reaction chamber 50.
  • the controlled sequence of PCR which is not self-evident in microfluidic systems, is described in US Pat Detection system 10 ensured by an over a compensation channel 38 with the reaction chamber 50 in fluid communication pressure chamber 60.
  • reaction chamber 50 Possibly in the reaction chamber 50 in the denaturation step at T ⁇ 96 ° C resulting gas bubbles press a small amount of the sample liquid in the pressure chamber 60, where it evaporates immediately because of the increased pressure chamber temperature. Due to the large volume of gas generated during the evaporation, a back pressure is rapidly generated in the pressure chamber 60 on the sample solution present in the reaction chamber 50, which greatly reduces the number and size of the gas bubbles.
  • the reaction chamber 50 and the pressure chamber 60 thus form a self-regulating subsystem for suppressing unwanted gas bubbles in the reaction chamber 50 and thereby ensure a well-controlled flow of the PCR.
  • a special implementation of the PCR is performed, which is referred to as real-time PCR, since it determines the amount of nucleic acid present in the sample solution while the reaction proceeds.
  • the Real Time PCR allows a more accurate determination of the concentration of nucleic acid in the sample solution as a pure end point determination at the end of a PCR.
  • a first predetermined number of cycles of the amplification reaction is first carried out, for example 15 cycles in order to increase the concentration of nucleic acid in the measurement range.
  • a defined volume of sample solution with amplified nucleic acids is transferred from the reaction chamber 50 into the detection chamber 40 (flow 36).
  • the sensor region 42 of a CMOS-based microchip is arranged, which has an array of functionalized test sites for electrochemical detection of the nucleic acids in the sample solution.
  • each of the test sites generates an electrical signal whose magnitude is a measure of the concentration of nucleic acid in the transferred detection volume.
  • a preferred electrochemical detection method is in the document DE 10 2011 056 606 B3 described, the disclosure of which is included in the present application in this respect.
  • a further PCR cycle is repeatedly carried out in each case to further increase the concentration of nucleic acid and after each cycle a defined volume of sample solution with amplified nucleic acids from the reaction chamber 50 is transferred into the detection chamber 40 and the concentration of nucleic acid is determined electrochemically. In this way, for example, a total of 30 to 40 PCR cycles can be performed. At the end of the procedure, a larger final volume of sample solution is drawn into the detection chamber 40 to determine the endpoint, and the concentration of nucleic acid is detected electrochemically.
  • a detection system 10 shows Fig. 2 schematically a detection system in the form of a microfluidic cartridge 100 for the detection of biomolecules in a sample solution, according to the associated with Fig. 1 explained principles is constructed.
  • Cartridge 100 is formed entirely of polycarbonate, except for the chemicals and microchip 142, for example by an additive manufacturing process or by injection molding.
  • the cartridge 100 is a disposable cartridge for cost-effective detection of desired nucleic acids, for example in an MRSA screening.
  • the cartridge 100 has a luer connection 110 to which, for example, a sample tube 90 with a swab sample 92 to be examined can be connected.
  • a microchannel system 120 that extends from a pumping device 200 via a detection chamber 140, a PCR chamber 150 with wax layer protected PCR chemicals 152, and a lysis chemicals storage area 170 to the luer port 110 , In other designs, lysis chemicals 172 are provided elsewhere, such as in the Luer port or sample tube 90.
  • the pump device is designed in the form of the double syringe 200 explained in more detail below and forms an integral part of the polycarbonate cartridge 100.
  • An outer chamber 204 of the double syringe 200 is pre-filled with a desired process liquid 202 in a storage-stable manner, so that the cartridge 100 is heated to the heating / Cooling device contains substantially all components required for the desired detection.
  • the cartridge contains a CMOS-based microchip 142 whose outline is in Fig. 2 indicated by dashed lines.
  • the microchip 142 has a in the Detection chamber 140 arranged sensor area 144 and a liquid-tight separated from the detection chamber contact area 146.
  • Fig. 3 schematically shows a cross section of the cartridge 100 in the region of the microchip 142 along the line III-III of Fig. 2 ,
  • the sensor region 144 of the microchip 142 contains an array of, for example, 109 functionalized test sites 130 in an octagonal arrangement for the electrochemical detection of the nucleic acids to be detected in the sample solution.
  • a suitable detection method is, for example, in the document DE 10 2011 056 606 B3 described.
  • each of the test sites 130 of the sensor area 144 is equipped with its own first-order sigma-delta modulator 132 for the AD conversion of the electrical signals generated at the test sites 130.
  • the sigma-delta modulators 132 provide as digitized signal a bitstream that is insensitive to noise and crosstalk, but which is responsible for complete AD conversion in accordance with per se known principles of the sigma-delta technique in a processing circuit 134 to produce the final Output signals is processed further.
  • the output signals can be picked up at the contact pads 136 of the contact region 146 of the microchip 142 and fed in a known manner to an evaluation and display unit.
  • the contact region 146 of the microchip 142 is separated in a liquid-tight manner, for example by means of a rubber ring 138 or a potting compound, from the detection chamber 140 with the sample solution contained therein.
  • the cartridge 100 is first connected to the sample system 90 via the Luer connection 110 and the valve 180 is opened, if not already open. Now, by a slight lifting 220 of the inner piston 216 of the double syringe 200 with an associated piston rod ( Fig. 7 (b) ) the through holes 218 between outer chamber and inner chamber 206 are released. By depressing 230 of the rubber ring 214, the supplied process liquid 202 is supplied to the sample 92 via the microchannel system 120, the process liquid in the storage area 170 receiving the lysis chemicals 172.
  • the depressed rubber ring 214 closes the through holes 218 from the outer chamber side, so that by a further axial movement of the inner piston 216 with the piston rod resulting from the lysis of the sample 92 sample solution into the microchannel system 120 of the cartridge 100 into the inner chamber 206 of the double syringe 200th can be withdrawn.
  • the channel system 120 including the PCR chamber 150 and the detection chamber 140 is filled with sample solution.
  • the pressure chamber 160 which is in fluid communication with the PCR chamber 150 via the compensation channel 122, remains substantially free of liquid.
  • valve 180 is closed to external sample system 90 to have a defined trapped fluid volume in the detection system for subsequent PCR.
  • the cartridge 100 can be used, for example, in a thermocycler, which provides heating / cooling means for the temperature steps of the PCR cycles.
  • a thermocycler which provides heating / cooling means for the temperature steps of the PCR cycles.
  • the PCR chemicals 152 are added to the sample solution and PCR is generated by generating the desired Cyclic temperature profile performed.
  • heating the pressure chamber 160 to a temperature T DK above the boiling point of the sample solution suppresses gas bubble formation in the PCR chamber 150, so that the PCR can proceed reliably and in a controlled manner.
  • a defined volume of sample solution containing amplified nucleic acids from the PCR chamber 150 is transferred into the detection chamber 140 by controlled lifting of the inner piston 216 of the double syringe.
  • the desired concentration levels of nucleic acid in the sample solution are determined by electrochemical detection of the nucleic acid oligomer hybridization events at the test sites 130 of the microchip 142, such as in U.S. Pat DE 10 2011 056 606 B3 described in more detail.
  • a further defined volume of further amplified sample solution is transferred from the PCR chamber 150 into the detection chamber 140 and the concentration of nucleic acid determined by successively raising the inner piston 216.
  • the PCR chamber 150 may have a volume of 30 ⁇ l, and after completion of the initial PCR cycles, a volume of 1 ⁇ l is transferred to the detection chamber 140 for 15 to 25 cycles after each PCR cycle.
  • a larger final volume of sample solution can be drawn into the detection chamber 140 and a concentration end point can be determined. It is understood that the concentration determination can also be carried out at longer intervals, for example after every second or every third PCR cycle.
  • FIG Fig. 4 A specific design of the cartridge 100 in the area of the detection chamber 140 and the PCR chamber 150 is shown in FIG Fig. 4 shown in perspective view.
  • the PCR chamber 150 is subdivided into a plurality of successively arranged, disk-shaped sub-chambers 154, of which only the last sub-chamber 154 is shown in the figure.
  • a supply channel 124 leads into the detection chamber 140, from which an outlet channel 126 extends to the double syringe 200.
  • CMOS-based microchip 142 The outline of the CMOS-based microchip 142 is again shown by dashed lines.
  • the sensor region 144 of the microchip 142 is arranged with the array of test sites 130 in the detection chamber 140 and separated in a liquid-tight manner from the contact region 146 with the contact pads 136 via a circumferential potting ring 139.
  • the self-regulating subsystem for suppression of gas bubbles in the PCR chamber is in Fig. 5 explained in more detail.
  • the principle described is not limited to use in a PCR, but can also be used in other temperature-controlled microfluidic processing processes in which a sample solution is to be kept free of gas bubbles near the normal boiling point of the sample solution.
  • FIG. 5 shows a section of a microfluidic device 400 for temperature-controlled processing of a sample solution 410, here the detection of nucleic acids described above.
  • the microfluidic device 400 includes a reaction chamber 450 in which the sample solution containing the nucleic acids is heatable by a heater / cooler 452 to a desired temperature near the normal boiling point of the sample solution 410.
  • the sample solution 410 is heated to about 96 ° C. in the denaturation step mentioned above.
  • gas bubbles can not only lead to large temperature differences in the sample solution in the reaction chamber 450, but also to squeeze sample solution out of the reaction chamber and to strong fluid oscillation in the microfluidic system.
  • several measures have been proposed, such as a special structural design of the reaction chamber, a surface treatment of the reaction chamber walls, a sealing of the reaction chamber under elevated pressure, a degassing of the sample solution and the addition of reagents high boiling point.
  • gas bubbles in the reaction chamber 450 are suppressed by a fluidic coupling of the reaction chamber 450 to a pressure chamber 460 which is liquid-free in normal operation and at a temperature above the normal boiling point of the sample solution 410.
  • the reaction chamber 450 is in fluid communication via a compensation channel 422 with a pressure chamber 460, which is heated by a heater 462 to a temperature T DK well above the normal boiling point of the sample solution.
  • gas bubble suppression is currently understood as follows: If, in the temperature-controlled processing of the biomolecules, such as the above-mentioned PCR denaturation step, gas bubbles 454 are formed in the reaction chamber 450, then a small sample liquid quantity 464 is forced through the equalization channel 422 into the pressure chamber 460 by the first forming gas bubbles, where due to the immediately evaporated in the pressure chamber 460 high temperature.
  • the volume of gas produced by the evaporation is 1000 to 2000 times greater than the volume of vaporized liquid, resulting in a large pressure increase in the pressure chamber 460, which in turn leads to an increase in pressure in the reaction chamber 450 and thus a reduction in the number and size of the gas bubbles leads and the formation of further gas bubbles counteracts.
  • the main effects, as currently understood, are the increase in vapor pressure and the compression of existing gas volumes.
  • the pressure chamber 460 can be easily integrated into a microfluidic system and requires the suppression of gas bubble formation does not involve complex designs of the reaction chamber, additions to the sample solution, or other expensive measures that either generally can prevent only nucleation of gas bubbles on the reactor walls or, as in the case of additions to the sample solution, can affect the PCR process ,
  • a microfluidic system such as the microchannel system 120, 140, 150, 170, 180 of FIG Fig. 2
  • a double syringe 200 according to the invention used, their construction and operating principle now with reference to Fig. 6 is explained in more detail.
  • the double syringe 200 contains two concentrically arranged cylinder chambers 204, 206, in each of which axially movable pistons 214, 216 are accommodated.
  • the end of the dual syringe 200 having the outlet port 210 is referred to as the distal end D of the syringe (s), the axially opposite end being the proximal end P.
  • the double syringe includes an outer cylindrical chamber 204 containing a process fluid reservoir 208 202 defined. Concentrically in the outer chamber 204, a cylindrical inner chamber 206 is arranged, which terminates at its distal end with the outer chamber 204.
  • the outlet port 210 of the double syringe is disposed in the distal cylinder top surface 212 of the inner chamber 206.
  • the two cylinder chambers 204, 206 are in fluid communication with each other via a series of through holes 218, which are formed circumferentially in the common cylinder wall 205 of the inner and outer chambers in the distal end region of the chambers 204, 206 at the same axial height.
  • an axially movable inner piston 216 is accommodated, which has on its proximal side a gripping element 222, which from the outside with a separate, adapted piston rod 240 (FIGS. Fig. 7 (b) ) can be gripped to axially move the inner piston 216 up and down (arrows 220).
  • a gripping element 222 for example, a cavity may also be formed in the proximal side of the inner piston 216, into which engages the correspondingly configured distal end of a piston rod.
  • the inner piston 216 When the inner piston 216 is located at the distal end of the inner chamber 206, the inner piston 216 closes the through-holes 218 from the side of the inner chamber with a circumferential seal 224, for example a rubber seal, so that no fluid flow from the reservoir 208 of the outer chamber 204 to the outlet opening 210 is possible.
  • This position of the inner piston 216 is therefore referred to as the closed position and is in Fig. 7 (a) shown.
  • the inner piston 216 moves slightly proximally, in the orientation of Fig. 6 thus raised, it releases the through holes 218 and allows fluid flow from the outer chamber 204 via the inner chamber 206 to the outlet 210. Such a position of the inner piston 216 is therefore referred to as the release position and is in Fig. 7 (b) shown.
  • an annular outer sealing piston surrounding the inner chamber 206 is accommodated, here in the form of a rubber ring 214, which is movable axially towards the distal end D of the outer chamber (arrows 230).
  • the rubber ring 214 closes the storage space 208 for the process fluid 202, as in Fig. 7 (a) shown.
  • the rubber ring 214 seals the through-holes 218 from the side of the outer chamber 204, thus allowing liquid to retract from the microfluidic system into the inner chamber 206 by axial movement of the inner piston 216 toward the proximal end P of the syringe.
  • a reservoir 226 may be provided for receiving chemicals.
  • the chemicals are initially fixed in the reservoir 226, but can by a release function, For example, a fusible wax cover or a magnetic particle cover layer may be released at a desired time.
  • Fig. 7 (a) In the initial state of Fig. 7 (a) is the inner piston 216 in its distal closed position and the rubber ring 214 in its proximal storage position.
  • the storage space 208 of the outer chamber 206 is prefilled with the desired process liquid 202 in a storage-stable manner.
  • the gripping element 222 of the inner piston 216 is gripped with an adapted piston rod 240 and inserted into the in Fig. 7 (b) raised release position in which the through holes 218, the fluid flow from the reservoir 208 of the outer chamber 204 to the outlet 210 release.
  • the process fluid 202 may be completely pumped into the microfluidic system (flow 242).
  • the double syringe can be used by moving the piston rod 240 axially up and down in the inner chamber 206 like an ordinary syringe and moving fluid in both directions.
  • fluid can be drawn from the microfluidic system into the inner chamber 206 (flow 244) by a further lifting, ie an axial movement of the inner piston 216 towards the proximal end.
  • the sample solution resulting from the lysis of the sample 92 is withdrawn from the sample tube 90 into the microchannel system of the cartridge 100 and ultimately into the inner chamber 206 of the double syringe.
  • successive small volumes of liquid are transferred from the PCR chamber 150 into the detection chamber 140 after each PCR cycle by controlled lifting of the piston rod 240.
  • the extension to the supply of three or more liquids is apparent from the following presentation.
  • the 2-fluid double syringe 300 of the Fig. 8 contains two concentrically arranged cylinder chambers 304, 306 and therefore constitutes a double syringe.
  • a cylindrical inner chamber 306 Concentrically in the outer chamber 304, a cylindrical inner chamber 306 is arranged, which terminates at its distal end with the outer chamber 304.
  • the outlet port 310 of the double syringe 300 is disposed in the distal cylinder top surface of the inner chamber 306.
  • Each group of through holes 318-1, 318-2 consists of a series of at the same axial height circumferentially formed in the common cylinder wall 305 of the inner and outer chamber through holes.
  • an axially movable inner piston 316 is accommodated, which has on its proximal side a gripping element 322 which can be gripped from the outside with a separate, adapted piston rod in order to move the inner piston 316 axially up and down (arrows 320 ).
  • a gripping element 322 which can be gripped from the outside with a separate, adapted piston rod in order to move the inner piston 316 axially up and down (arrows 320 ).
  • a cavity in the proximal side of the inner piston 316 may be formed, in which engages the correspondingly configured distal end of a piston rod.
  • the inner piston 316 When the inner piston 316 is moved slightly proximally, it initially releases only the first group of through-holes 318-1 and allows fluid flow of the first fluid 302-1 from the outer chamber 304 via the inner chamber 306 to the outlet port 310. This position of the inner piston 316 is referred to as the first release position and is in Fig. 8 (b) illustrated. By a further proximal movement of the inner piston 316 reaches the second release position in which the second group of through holes 318-2 is released from the inner chamber side (see Fig. 8 (d) ).
  • the inner chamber 306 are housed surrounding axially spaced rubber rings 314-1, 314-2 which are each axially movable toward the distal end D of the outer chamber (arrows 330-1, 330-2).
  • the rubber rings 314-1, 314-2 respectively close off a reservoir for one of the deliverable fluids 302-1, 302-2, as in FIG Fig. 8 (a) shown.
  • the first rubber ring 314-1 closes the first group of through-holes 318-1 (see FIG Fig. 8 (c) ) while the second rubber ring 314-2 closes the second group of through-holes 318-2 in its distal closed position (see FIG Fig. 8 (e) ).
  • the inner piston 316 is in its first or second release position, then, as a result of the movement of the two rubber rings 314-1, 314-2 or only the second rubber ring 314-2 towards the distal end, they can be successively introduced into the first or second reservoir Liquid is forced into the inner chamber 306 via the first and second groups of through-holes 318-1, 318-2, respectively. If both rubber rings 314-1, 314-2 brought into their distal closed position, all through holes are closed and the liquid can by a proximal movement of the inner piston 316 are withdrawn from the microfluidic system into the inner chamber 306.
  • the gripping element 322 of the inner piston 316 is gripped with an adapted piston rod and moved into the first release position (FIG. Fig. 8 (b) ) raised.
  • the first process fluid 302-1 can be completely pumped into the microfluidic system (flow direction 342). Because of the incompressibility of the fluids, the uppermost of the n rubber rings, it is sufficient to actively press the rubber ring 314-2 distally.
  • the pressure exerted is transferred to the first rubber ring 314-1 via the incompressible second process fluid 302-2, so that the two rubber rings move distally distally and the volume of the reservoir 308-2 moves upon supply of the first process fluid 302-1 remains unchanged (cf. Figs. 8 (a) and 8 (c) ).
  • the first rubber ring 314-1 and the position of the first group of through-holes 318-1 in the cylinder wall 305 are matched to each other such that the first rubber ring 314-1 closes the first group of through-holes 318-1 from the outer chamber side in its distal closed position , as in Fig. 8 (c) shown.
  • the inner piston 316 further in the second release position ( Fig. 8 (d) ), and by squeezing the second rubber ring 314-2, the second process liquid 302-2 is completely pumped into the microfluidic system (flow direction 344).
  • the second rubber ring 314-2 and the position of the second group of passage openings 318-2 in the cylinder wall 305 are matched to one another such that the second rubber ring 314-2 in its distal closure position is the second Group of through-holes 318-1 from the outer chamber side closes, as in Fig. 8 (e) shown.
  • the double syringe 300 can be used by moving the inner piston 316 axially up and down with the piston rod like a conventional syringe and moving fluid in both directions.
  • liquid can be drawn from the microfluidic system into the inner chamber 306 (flow direction 246) by a further lifting, ie an axial movement of the inner piston 316 towards the proximal end.
  • the first n-1 annular piston need not necessarily be sealing, for example, the annular piston 314-1 of the double syringe 300 with a little clearance to the cylinder walls of the inner and outer chamber and only the uppermost annular piston 314-2 sealing ,
  • n ring pistons in their distal closure position together close the one or more groups of through-holes from the outer chamber side, so that Retracting liquid from the microfluidic system into the inner chamber is made possible by an axial movement of the inner piston towards the proximal end when all n annular pistons are brought into their distal closed position.

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Claims (13)

  1. Double seringue, destinée à amener un liquide dans un système micro-fluidique, comprenant
    - un compartiment extérieur cylindrique, qui définit un espace de réserve, destiné à recevoir le liquide qui doit être amené,
    - un compartiment intérieur cylindrique, placé à l'intérieur du compartiment extérieur, qui dans sa surface de recouvrement distale cylindrique comporte un orifice de sortie, destiné à être assemblé avec un système micro-fluidique et qui par l'intermédiaire de perçages de passage est en liaison fluidique avec le compartiment extérieur,
    - un piston intérieur, déplaçable en direction axiale dans le compartiment intérieur, qui dans une position de fermeture ferme les perçages de passage vers le côté du compartiment intérieur et qui dans une position de libération libère les perçages de passage et permet un flux de fluide du compartiment extérieur dans le compartiment intérieur et
    - un piston annulaire, mobile en direction axiale dans le compartiment extérieur, en direction de l'extrémité distale du compartiment extérieur,
    i) qui dans une position de réserve proximale, ferme l'espace de réserve pour le liquide qui doit être amené,
    ii) par le déplacement duquel en direction de l'extrémité distale, un liquide présent dans l'espace de réserve est poussé par l'intermédiaire des perçages de passage dans le compartiment intérieur, lorsque le piston intérieur se trouve dans sa position de libération et
    iii) qui dans une position de fermeture distale ferme les perçages de passage à partir du côté du compartiment extérieur et permet de ce fait un retrait vers l'arrière de liquide hors du système micro-fluidique dans le compartiment intérieur, par un déplacement axial du piston intérieur en direction de l'extrémité proximale.
  2. Double seringue selon la revendication 1, caractérisée en ce que sur son côté proximal, le piston intérieur comporte un élément de préhension qui est susceptible d'être saisi par une tige de piston correspondante, pour le déplacement axial du piston intérieur.
  3. Double seringue selon la revendication 1, caractérisée en ce que sur son côté proximal, le piston intérieur comporte une cavité dans laquelle l'extrémité distale d'une tige de piston correspondante peut s'engager, pour le déplacement axial du piston intérieur.
  4. Double seringue selon l'une quelconque des revendications 1 à 3, caractérisée en ce que le piston intérieur est muni d'un joint en caoutchouc, destiné à fermer les perçages de passage.
  5. Double seringue selon l'une quelconque des revendications 1 à 4, caractérisée en ce que la position de fermeture du piston intérieur se situe sur l'extrémité distale du compartiment intérieur cylindrique et en ce que dans celle-ci, le piston intérieur jouxte sensiblement la surface de recouvrement cylindrique du compartiment intérieur.
  6. Double seringue selon l'une quelconque des revendications 1 à 5, caractérisée en ce que le piston annulaire du compartiment extérieur est formé par une bague en caoutchouc.
  7. Double seringue selon l'une quelconque des revendications 1 à 6, caractérisée en ce que la position de fermeture du piston annulaire se situe sur l'extrémité distale du compartiment extérieur cylindrique et en ce que dans cette dernière, le piston annulaire jouxte sensiblement la surface de recouvrement cylindrique du compartiment extérieur.
  8. Double seringue selon l'une quelconque des revendications 1 à 7, caractérisée en ce que les perçages de passage sont conçus dans la paroi cylindrique commune du compartiment intérieur cylindrique et du compartiment extérieur cylindrique, de préférence dans la zone d'extrémité distale de la paroi cylindrique, sur la même hauteur axiale, en périphérie autour de la paroi cylindrique.
  9. Double seringue selon l'une quelconque des revendications 1 à 8, caractérisée en ce que l'espace de réserve est pré-rempli d'un liquide de processus.
  10. Double seringue selon l'une quelconque des revendications 1 à 9, caractérisée en ce que dans le côté proximal du piston intérieur est prévu un espace de réserve, destiné à recevoir des produits chimiques, qui est rempli notamment de produits chimiques souhaités et d'une fonction de libération.
  11. Double seringue, destinée à amener ≥ 2 liquides dans un système micro-fluidique, comprenant
    - un compartiment extérieur cylindrique, qui définit n espaces de réserve destinés à recevoir les liquides qui doivent être amenés,
    - un compartiment intérieur cylindrique, placé à l'intérieur du compartiment extérieur, qui sur son extrémité distale comporte un orifice de sortie, destiné à être assemblé avec le système micro-fluidique et qui par l'intermédiaire d'un ou de plusieurs groupes de perçages de passage, écartés en direction axiale est en liaison fluidique avec le compartiment extérieur,
    - un piston intérieur, déplaçable en direction axiale dans le compartiment intérieur, qui dans une position de fermeture, ferme l'un ou les plusieurs groupes de perçages de passage sur le côté du compartiment intérieur et qui dans une ou dans plusieurs positions de libération, permet un flux de fluide du compartiment extérieur dans le compartiment intérieur et
    - n pistons annulaires, déplaçables dans le compartiment extérieur, en direction axiale vers l'extrémité distale du compartiment extérieur,
    i) qui respectivement dans une position de réserve proximale, ferment un espace de réserve pour l'un des liquides susceptibles d'être amenés,
    ii) dont le déplacement vers l'extrémité distale pousse successivement un liquide présent dans le k ème espace de réserve par l'intermédiaire de perçages de passage dans le compartiment intérieur, lorsque le piston intérieur se trouve dans une position de libération correspondante, k étant = 1, ... n,
    iii) dans leur position de fermeture, les n pistons annulaires ferment ensemble l'un ou les plusieurs groupes de perçages de passage, à partir du côté du compartiment extérieur, de telle sorte
    iv) qu'un retrait vers l'arrière de liquide hors du système micro-fluidique dans le compartiment intérieur par un déplacement axial du piston intérieur vers l'extrémité proximale soit possible, lorsque tous les n pistons annulaires sont amenés dans leur position de fermeture distale.
  12. Double seringue destinée à amener ≥ 2 liquides dans un système micro-fluidique selon la revendication 11, caractérisée en ce que
    - le compartiment intérieur est en liaison fluidique avec le compartiment extérieur par l'intermédiaire de n groupes de perçages de passage, écartés en direction axiale,
    - dans une position de fermeture, le piston intérieur ferme les n groupes de perçages de passage sur le côté du compartiment intérieur et dans n positions de libération écartées en direction axiale, libère successivement un nombre croissant 1, ..., n de groupes de perçages de passage et permet un flux de fluide du compartiment extérieur dans le compartiment intérieur,
    - par le déplacement des pistons annulaires déplaçables dans le compartiment extérieur vers l'extrémité distale, un liquide présent dans le k ème espace de réserve est poussé successivement par l'intermédiaire du k ème groupe de perçages de passage dans le compartiment intérieur lorsque le piston intérieur se trouve dans la k ème position de libération, k étant = 1, ... n, et
    - dans une position de fermeture distale, le k ème piston annulaire ferme le k ème groupe de perçages de passage, à partir du côté du compartiment extérieur.
  13. Cartouche micro-fluidique, destinée à déceler des biomolécules dans une solution échantillon, pourvue d'un système de micro-canaux et d'une double seringue selon l'une quelconque des revendications 1 à 12, destinée à amener un liquide de processus dans le système de micro-canaux.
EP16708075.3A 2015-02-20 2016-02-16 Double séringue pour amener un liquide dans un système micro-fluidique Active EP3259064B1 (fr)

Applications Claiming Priority (2)

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DE102015001999.6A DE102015001999B3 (de) 2015-02-20 2015-02-20 Doppelspritze für die Zuführung einer Flüssigkeit in ein Mikrofluidiksystem
PCT/EP2016/000258 WO2016131537A1 (fr) 2015-02-20 2016-02-16 Double séringue pour amener un liquide dans un système micro-fluidique

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CN113266702A (zh) * 2020-02-14 2021-08-17 安行生物技术有限公司 控制流体的装置和方法
US20230105360A1 (en) * 2020-03-09 2023-04-06 Nuclein, Llc Apparatus and methods for molecular diagnostics
EP4273430A4 (fr) 2020-12-31 2024-12-25 Everlast Healthcare Limited Mécanisme à piston, mécanisme de commande de fluide et son application

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US7025323B2 (en) * 2001-09-21 2006-04-11 The Regents Of The University Of California Low power integrated pumping and valving arrays for microfluidic systems
DE10149684B4 (de) * 2001-10-09 2005-02-17 Clondiag Chip Technologies Gmbh Vorrichtung zur Halterung eines Substanzbibliothekenträgers
ATE376882T1 (de) * 2003-07-04 2007-11-15 November Ag Molekulare Medizin Verwendung eines einwegbehälters, mikrofluidische vorrichtung und verfahren zur bearbeitung von molekülen
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DE102010043030A1 (de) 2010-10-28 2012-05-03 Robert Bosch Gmbh Mikrofluidische Vorrichtung und mikrofluidisches Verfahren zur Verarbeitung von Biopartikeln
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WO2016131537A1 (fr) 2016-08-25
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