Patent Application
20120184046
This invention relates generally to the manufacture of complex layered materials and devices. More particularly, the present invention relates to methods of selectively bonding two surfaces by selectively modifying or coating at least one surface prior to bonding to reduce, or prevent, the bonding in the selected areas. The field of this invention also extends to the manufacture of complex polymeric materials and devices, in particular those for use in microfluidic applications.
This invention also relates to structures, devices and methods of manufacture for optical imaging in microfluidic devices using porous material inside detection flow cells.
Many industries have moved to using layered materials to take advantage of the increased material characteristics and functionality provided by such composite structures. In some instances the fabrication of devices from layered materials can simplify the manufacturing process by forming 3-dimensional (3D) components by stacking and bonding multiple layers that have been machined or processed separately. In the field of microfluidics the layering of materials is particularly important to seal the microstructures.
In polymer microfluidic fabrication many of the manufacturing approaches are limited to creating 2-dimensional or 2½-dimensional structures. The most common of these approaches use either computer numerical control (CNC) micromilling, injection-moulding or hot embossing, which can generate only very limited feature complexity. The fabrication of complex 3-dimensional parts typically requires the assembly of several separately machined parts. However, these are often serial fabrication processes that have alignment challenges when assembling micro-parts which lead to further labour-intensive processes with relatively low throughput and high associated production costs.
Another recent approach to the fabrication of polymeric microfluidic devices is the stacking, aligning and bonding of several layers of thin, already fabricated films. This layered approach allows the use of relatively simple 2-dimensional manufacturing techniques (such as embossing, die cutting, and laser processing) as well as established bonding technologies to create complex three-dimensional materials or devices. Such a 3D design approach is especially suited to high-volume manufacturing using reel-to-reel processing as described recently by Mehalso (Robert Mehalso, “The Microsystems road in the USA” Mstnews, Volume 4/02, pgs 6-8 (2002)), Schuenemann et al. (Matthias Schuenemann, David Thomson, Micah Atkin, Sebastiaan Gars, Abdiraham Yussuf, Matthew Solomon, Jason Hayes, Erol Harvey, “Packaging of Disposable Chips for Bioanalytical Applications”, IEEE Electronic Components & Technology Conference, Nevada, USA 2004), and WO 2007/085043.
In polymer microfluidics, bonding represents a particularly difficult problem due to the requirements of maintaining the integrity of the microstructures while forming a good seal.
Bonding techniques may be broadly classified into two categories; Area bonding in which the entire surfaces of two substrates are bonded together, and Selective bonding in which selective regions on the surfaces are bonded together. Both techniques may be applied to microfluidic bonding. Typically selective bonding is the more expensive technique to implement in production but the spatial control of the bonding seal may be greater, reducing the risk of interfering with microstructures.
Adhesive bonding is typically the most common method used in polymer microfluidics. This method requires another material to act as a linker to bond two surfaces together. Typical adhesives include: cyanoacrylates, silicones, epoxies, and acrylic based materials. In manufacturing setting adhesives can be easily coated over an entire surface by sprays, wire bars, doctor blades, rollers, or laid down as a sheet or tape. Furthermore the lifetime performance, toxicity and surface interactions are all important considerations particularly for microfluidic devices in which the surface to volume ratios are so large. These are often causes of failure in these devices where the adhesive is exposed to the microfluidic channel. Therefore in many microfluidic applications it is critical to choose a compatible adhesive or enable selective adhesive control. A limited set of adhesives can be selectively deposited by printing techniques, such as the use of hot melt adhesive, or with patterned adhesive sheets or tapes. However it can be difficult to selectively deposit adhesives in a volume manufacturing setting due the select availability of suitable adhesives and deposition techniques. Some of the many issues include the adhesive viscosity requirement, the adhesive's lifetime prior to bonding, speed of deposition and deposition control.
The Diffusion method is also commonly employed in polymer microfluidics as it requires does not require the addition of any chemicals that might adversely impact device performance. U.S. Pat. No. 5,882,465 describes such a method whilst bonding under vacuum pressure to reduce the chance of bubble formation. This common batch-based technique involves applying pressure and temperature whilst bringing the substrate surfaces together and allowing time for the molecular chains from each material to slowly diffuse into one another. Typically this requires similar materials having molecular chains with sufficient mobility. Although many layers can be bonded at once care needs to be taken with voids weakening bonding layers and the applied pressures deforming structures. From a manufacturing view the process requires relatively long processing times which limits the throughput capability.
Surface modification by techniques such Plasma, corona, or UV assisted bonding have been described in the literature and they involve changing the surface chemical groups to improve bonding. Typically the exposure of a polymer in an oxygen atmosphere by one of these techniques can lead to an increase in the surface oxygen groups, which increases the surface energy and enhances bonding for many substrates. Other gases and liquids on the surface can be exposure to produce other functional surface groups. Many of the reaction pathways created by these exposure techniques involve unstable free radical species. However the suitability of these techniques has only been demonstrated for a few materials.
In limited cases selective bonding can been achieved by surface modification if masking techniques are used, ensuring the exposed areas are limited to the bonding areas. However this can be difficult to implement in a high speed production environment and still maintain the tight tolerances required for microstructured devices.
Solvent assisted bonding uses solvents to swell the polymer surfaces and increase the chain mobility to allow the two surfaces to diffuse into one another. Generally the main problem with this technique is the difficulty of handling the solvents in the production environment. Furthermore, for fluidic devices the solvent residues can provide a source of contamination, and the solvent may deform the microstructures. A process for combining a weak solvent with heat activated bonding is described in US Patent Application 2008178987.
Transmission laser welding operates by one material being transparent to and the other material being an absorber to the irradiated laser wavelength. This allows the laser beam to selectively heat between the two materials producing localised welding when the heat goes above the glass transition temperature. For integration into the production environment, the main limitations are processing times, and limitation of compatible materials and number of layers that can be processed.
Reverse conduction welding operates in a similar manner to transmission layer welding except that the heat is generated by laser absorption at a backplane. The polymer films clamped above the absorbing layer conduct the heat from its surface and locally melt. Due to the uniform heat conduction within the polymers which limits spatial resolution, the technique is only suitable for thin films and relatively large structures.
High frequency or dielectric heating is a technique that can bond polar materials by passing an AC current through them. This method can be effective for bonding materials that would normally degrade near their softening point. This is because the heat is generated uniformly in the material rather than at the surface and then conducted inwards. However for microstructures, this can introduce problems due the non specific heating causing deformation.
Ultrasonic welding depends on vibration energy being transmitted through the materials. At the interface of the two materials the vibrationary energy is translated into heat. Features can be used to focus the energy, and with careful energy control and geometry design around structured parts a good seal can be achieved without deforming the remaining material. Due to these geometric constraints for bonding, ultrasonic sealing is limited in terms of its application to microfluidics.
The deposition of specific energy absorbing materials in the proximity of the join can be also be used to induce localised melting and therefore selective bonding when irradiated by the appropriate energy sources. Energy absorbers include thin film metals, Clearweld™, polyaniline, polypyrrole, polyalkylthiophenes, metallic nanoparticles, magnetic and paramagnetic particles and other appropriately doped materials. Energy sources include electromagnetic, Microwave, UV/Visible, and Infrared radiation. For sealing microstructures the effectiveness is typically dependant limited by the deposition technique and evenly controlling the energy absorbed.
Lamination is a popular technique for joining plastic films by bringing the materials together with one or more of the films having an adhesion layer. This adhesion layer may be an adhesive as described above, or a polymer with a lower glass transition temperature that will flow under temperature and pressure to bond to the other surface. These methods are widely used in the printing and packaging industries on reel to reel systems and have been applied to microfluidic devices (A. Schwarz F. Bianchi R. Ferrigno F. Reymond H. H. Girault J. S. Rossier, Microchannels Networks for Electrophoresis Separations, 20 Electrophoresis. 727(1999)). In similar manner the lamination of layers where at least one of those layers is an adhesive layer (such as a pressure sensitive adhesive) is commonly used in microfluidics (Robert Mehalso, “The Microsystems road in the USA” Mstnews, Volume 4/02, pgs 6-8 (2002)). However these lamination methods are area bonding techniques that bond all the surfaces which are in contact. For many microstructures in polymeric devices this is further complicated by the deformation of the structure during the bonding process. If adjacent surfaces are in contact during the applied pressure then a bond may form. The use of adhesive tapes for microfluidics is further complicated by chemical or biochemical incompatibility with many assays, and the dimensional limitations provided by the machining processes of these tapes.
Lamination and other area bonding techniques are advantageous to simplify manufacturing, allowing both speed and cost improvements (WO 2007/085043, the entire contents of which are incorporated herein by reference). However, with all these area bonding techniques a problem arises where a bond is not required, or required at a different strength, in a selective area between two surfaces in contact with one another. In many cases selective bonding is not an option due to material compatibility, cost, speed and dimensional constraints. What is needed for microfluidic production is a technique that allows the selective deactivation of surfaces that is compatible with bonding techniques suitable for mass production.
The demand for rapid and easy to operate point of care in-vitro assays continues to rapidly grow. The major need is for rapid, simple (preferably single-step) reliable assays that detect specific analytes and can be easily performed outside of the laboratory setting, be it by patients at home, in the doctor's office, or at any remote location.
“Dipstick,” lateral flow,” and “flow through” format systems are typical point of care systems in use today. They are designed for rapid on-site detection of various analytes. The dipstick type of assays and devices are exemplified in U.S. Pat. Nos. 4,059,407; 5,275,785; 5,504,013; 5,602,040; 5,622,871; and 5,656,503.
A dipstick point of care device typically consists of a strip of porous material made up of three contiguous parts—a sample receiving end, a reagent zone, and a reaction zone. Different materials, usually porous, are used for the different zones, but are typically combined to form a single strip or dipstick.
Either the liquid sample is applied to the sample zone, or the sample zone is dipped into the liquid sample. The liquid sample then wicks along the porous strip into the reagent zone where the analyte binds to a reagent, already pre-incorporated into the strip in the reagent zone, thus forming a complex. The complex is usually either an antibody/antigen pair or a receptor/ligand that creates a label. The labeled complex continues its wicked migration into the reaction zone where the complex binds to another specific binding partner and is immobilized. The result provides some kind of visual readout.
Typically lateral flow devices use porous material with a linear construction similar to that of dipsticks, incorporating the three sample, reagent release zone and reaction zones. Rather than vertically wicking the sample up the dipstick, lateral flow devices flow across the porous material. Examples of assays and devices using the lateral flow format can be found in U.S. Pat. Nos. 4,943,522, 5,075,078; 5,096,837; 5,229,073; 5,354,692; 6,316,205; and 6,368,876, the contents of which are incorporated herein by reference.
Similar components are sometimes often in both flow-through and lateral flow devices. The key difference is the components in such a flow through device, which are stacked one on top of the other to enable a unilateral downward flow. In most cases in such a flow-through device, the sample application pad sits over and in direct contact the conjugate pad, which sits on the analytical membrane, under which lies an absorbent pad.
In other examples of flow through assays the fluid is gravity fed through a column of frits with separator porous elements and optical analysis, WO 2008/145722. As with other afore mentioned lateral and vertical flow devices the effect of capillary action or gravity driven flow is limited to relatively simple protocols as multiple flows from different sources and complex flow profiles, such as backwashing, are not feasible.
As can be seen from the above descriptions of typical analyte detection devices, the sample receiving area, reagent area, reaction area or analytical membrane, and the absorbant material may be all made from porous materials, such as porous polymeric materials. Limitations of such systems include the reliance on capillary or gravity flow for fluid movement, which inherently causes reproducibility issues with regards to flow rate and limitations in terms of suitability of assay protocols. These capillary and gravity flow devices are limited in terms of performing only simple one-step assays; they provide imprecise handling of fluid volumes which affects the overall reproducibility; they are restricted in terms of the maximum volume they can use and therefore limits the sensitivity; they are susceptible to matrix effects obstructing pores; and they typically provide a qualitative or semi-quantitative response [Analytical and Bioanalytical Chemistry, Volume 393, Number 2, January 2009, pp. 569-582(14)].
Microfluidics techniques have been developed that provide accurate control of flow in small structures. These developments have been brought about by the advantages that miniaturization has to offer. In particular, performance improvements can be achieved over traditional laboratory equipment in terms of automation, reproducibility, speed, cost and size. This rapidly growing field includes micro total analytical systems (μTAS), or “lab on a chip” devices. Much of this early work was performed on silicon or glass substrates using established techniques developed in the 70's and 80's for the semiconductor industries. There have been many different pumping and valving strategies that have been integrated into miniaturized devices.
Critical to the usability of microfluidic devices in many applications is the ability to analyze the characteristics of the fluids contained within the microstructures. Optical detection strategies remain one of the most common methods used to measure these characteristics in microfluidic devices. Such optical detection strategies encompass absorption, transmission and luminescence (commonly chemiluminescence and fluorescence) based measurements.
Most of these difficulties in optical measurement within microstructures arise from the tight dimensional constraints, reduced path lengths, and reduced fluid volumes leading to much smaller signal responses. Methods to increase sensitivity and dynamic range often involve increasing the amount of sample volume and or the amount reporter reagent. A porous solid phase gives a relatively high surface area for binding in comparison to binding to the walls of a capillary, well, or chamber in a microfluidic device. Such porous materials are typically used to bind the analytes of interest and allow removal of unwanted reagents in affinity chromatography, such as with immunoassays and DNA hybridisation.
One of the advantages of microfluidics is that the smaller volumes of fluid typically result in a speed improvement in detection due to the reduction in distance between the analyte in solution and the sensor surface. However a problematic aspect of microfluidic device manufacture is the increase in cost associated with the manufacturing processes required to achieve smaller dimensions and their associated tolerances. Polymers have been used as a cheaper alternative to glass and silicon for manufacturing consumable devices, especially since the 1940's and have been used for mass producing complex materials and devices for instrumentation since the early to mid 1990's. However for polymer device fabrication it is generally known that as the dimensions of a feature on a device decreases in size and the tolerance required, the cost and difficulty in implementing in a mass manufacturing environment increases greatly. This is particularly problematic in microfluidics where the tolerance requirements are often much less than 100 micron. Examples of manufacturing methods for feature formation in microfluidic devices can be generally classified into two categories. The first is using direct machining methods in which the pattern of desired features is created directly on the surface of a stratum made of a suitable material. These methods include micromilling, laser based lithography and beam scanning, plasma etching, wet chemical UV lithography using photoresists, soft lithography, x-ray lithography and print-head deposition. The second methodology involves processes that use a master template to form the desired pattern. These feature replication processes include, soft lithography, stamping, embossing, compression molding, thermoforming, injection molding and reaction injection molding.
This invention combines the fluid manipulating advantages of microfluidics with porous structures for improved methods of detection in affinity chromatography with a method that is cost effective for mass manufacturing.
All of the processes described above are applicable to the process according to the present invention described herein.
The reference to any prior art in this specification is not, and should not be taken as, an acknowledgement or any form of suggestion that the prior art forms part of the common general knowledge.
This invention relates generally to the manufacture of complex layered materials and devices, and in particular to the manufacture of microfluidic devices. The invention overcomes the limitations described for the bonding of structured layers by providing a method for selectively reducing the bonding of materials.
A bond-reducing material is used to either fully or partially prevent a bond forming in a spatially defined location, and may be used improve the surface characteristics in a microstructure.
In one aspect, the present invention provides a method for forming a spatially defined bond between a first surface and a second surface, the method comprising the steps of (i) printing a bond-reducing material to an area on the first surface, and (ii) contacting the first surface and the second surface under conditions allowing the first surface to bond to the second surface, wherein the bond-reducing material substantially prevents or otherwise interferes with the formation of a bond between the first surface and the second surface about the area to which the bond-reducing material is applied, wherein the structure resulting from bonding first surface and the second surface is a microfluidic device.
In one embodiment, the bond-reducing material is printed by a process selected from the group consisting of: Microspotting (contact or non-contact); Contact printing; Screen printing; Syringe or ink-jet delivery; Lithography; robotic placement of dried or liquid chemicals; Letterpress, Gravure, flexographic and other such printing methods; contact mask based deposition methods; Laser based deposition or surface modification techniques; and thermal transfer methods, such as with laser, hot stamping, and thermal ribbon printers.
In one embodiment, the bond-reducing material is selected from the group consisting of an ink: A) Colorants (including pigments, toners, and dyes) that provide colour contrast. B) Vehicles, or varnishes, that bind to the printed surface and may act as carriers for any colorants during the printing operation. C) Additives that influence the printability, film characteristics, drying speed, or end-use properties, such as the inclusion of chemical moieties for bond reduction. D) Solvents, which may help in formation of the vehicles, in reducing ink viscosity, adjusting drying properties, or resin compatibility.
In one embodiment, the bond-reducing material is a solid film or foil, powder, high-viscosity paste, gel, or a low-viscosity liquid.
In one embodiment, the first surface is bonded to the second surface by a method selected from the group consisting of laser welding, diffusion bonding, surface modified chemical bonding, solvent assisted bonding, thermal laminating, chemical covalent or charged surface group bonding, mechanical interlocking, ultrasonic welding, die-electric bonding, microwave bonding, electrostatic or magnetic attraction, and adhesive bonding.
In one embodiment, the bond-reducing material is at least partially removed by a method selected from the group consisting of evaporation, absorption, chemical reaction or the application of mechanical force, air or liquid pressure.
In one embodiment, a composite structure formed by the spatially-selective bonding a first structure to a second structure, the composite structure having in one area a cross-sectional arrangement comprising the first structure, a bond-reducing material, and the second structure; and in another area the first structure, a bond-forming material and the second structure.
In a further aspect, the present invention provides a composite structure wherein the first structure or the second structure are materials selected from the group consisting of: polyolefin; Cyclo Olefin Polymer; polypropylene; polyethylene; low density polyethylene; high density polyethylene; polymethyl-methacrylate; polycarbonate; polyethylene terephthalate; polyethylene terephtalate glycol; polybutylene terephtalate; polystyrene; polyimide; polyetherimide; acrylonitrile butadiene styrene; polyurethane; polydimethylsiloxane; cellulose acetate; polyamide; polyether ether ketone; polyvinylchloride; polyvinylidene chloride; polyvinylidene fluoride; polymethylpentene; polysulfone; polytetrafluoroethylene; polyoxide methylene; nitrocellulose, nylons, acrylics, acetates, polyacrylamides, latex or silica particles, glass fibres or combinations thereof.
In one embodiment, the composite structure is produced by a method described herein.
In a further aspect the present invention provides a microfluidic device comprising a composite structure described herein.
In a further aspect the present invention provides a substantially planar microfluidic device for the affinity chromotographic analysis of a liquid analyte, the device comprising a substantially larger detection flow cell than the connecting microfluidic channels, the detection flow cell disposed substantially perpendicular to the plane of the device, the flow cell comprising (i) a liquid entry aperture (ii) a porous region and (iii) a liquid exit aperture, wherein in use the analyte flows from the liquid entry aperture, through the porous region and exits the flow cell via the liquid exit aperture.
In one embodiment the substantially larger detection flow cell is disposed at an angle of about 45, 50, 55, 60, 65, 70, 75, 80, 85, or 90 degrees relative to the plane of the device.
In one embodiment, the substantially larger detection flow cell is disposed at an angle of about 90 degrees relative to the plane of the device.
In one embodiment, the detection flow cell is capable of sustaining a maximum flow rate of 1000 micro litres per minute.
In one embodiment, the detection flow cell has a length of 10 micron to 10 millimetres.
In one embodiment, the detection flow cell has a width of 100 micron to 10 millimetres.
In one embodiment, the detection flow cell is substantially of cylindrical or rectangular shaped.
In one embodiment, the detection flow cell comprises a polymer frit.
In one embodiment, the detection flow cell comprises an affinity ligand.
In one embodiment, the detection flow cell comprises an affinity chromatographic resin.
In one embodiment, the device having a size and detection flow cell layout compatible with standard microtiter plate based systems.
In one embodiment, the device having a multi-layer laminate comprising microfluidic structures.
In one embodiment a device substantially as described in the drawings
In a further aspect the present invention a microfluidic affinity chromatographic method is provided, the method comprising (i) introducing an analyte into the detection flow cell of a device according to any one of claims 12 to 21 under conditions allowing the binding of a target molecule in the analyte to an affinity ligand and (ii) detecting the presence or absence of a bound target molecule.
a and 1b illustrates a bond-reducing material coated onto one substrate prior to and after bonding.
a and 4b illustrates a bond-reducing material containing magnetic properties with and without an applied magnetic field.
It is convenient to describe the invention herein in relation to particularly preferred embodiments relating to microfluidic devices. However, the invention is applicable to a wide range of situations and products and it is to be appreciated that other constructions and arrangements are also considered as falling within the scope of the invention. Various modifications, alterations, variations and or additions to the construction and arrangements described herein are also considered as falling within the ambit and scope of the present invention.
The invention overcomes the limitations described for the bonding of structured layers by providing a method for selectively reducing the bonding of materials. In the context of this invention a bond-reducing material is defined as a material that is used to reduce the strength of a bond between two surfaces, or prevent a bond that would have otherwise occurred between two surfaces. The bond reducing material may be applied prior to or during the bonding process. In its most generic form, the invention uses a bonding technique in combination with a printing method to modify or cover at least one portion of a surface with a bond-reducing material to either fully or partially prevent localised bonding. The structuring process may act upon the layers either before or after the bonding of the layers.
The advantages of this invention for the bonding of microfluidics are numerous. Firstly it provides a simplified manufacturing method suitable for high-throughput production. It also enables a greater spatial control over the bonding process by using known printing methods to provide controlled bonding areas. There is also the added advantage that numerous spatial and area bonding techniques can be used that would otherwise be unsuitable due to their spatial resolutions or incompatibility with the microfluidic application.
The function of the bond-reducing material is to either fully or partially prevent the bond forming in a spatially defined location and or improve the surface characteristics in a microstructure. The bond-reducing material may effect either a permanent change in the surface, or a transient change that is present during the bonding process. In one embodiment the bond-reducing material comprises a permanent coating. In another embodiment bond-reducing material comprises a transient volatile component, or non-volatile component that is physically removed after the bonding process. In one embodiment the removal of the transient component occurs by evaporation, absorption, chemical reaction or the application of mechanical force, air or liquid pressure either during manufacture or during the operation of the device.
In one embodiment the bond-reducing material comprises one or more ink components, such as A) Colorants (including pigments, toners, and dyes) that provide colour contrast. B) Vehicles, or varnishes, that bind to the printed surface and may act as carriers for any colorants during the printing operation. C) Additives that influence the printability, film characteristics, drying speed, or end-use properties, such as the inclusion of chemical moieties for bond reduction. D) Solvents, which may help in formation of the vehicles, in reducing ink viscosity, adjusting drying properties, or resin compatibility. The bond-reducing material may be a solid film or foil, powder, high-viscosity paste, gel, or a low-viscosity liquid. The various drying, curing or attachment methods may include heating, oxidizing, UV cross-linking, evaporating, penetrating, precipitating, polymerizing, reactive, including radiation-cured, gelling, cold-setting or quick-setting, and thermosetting.
In a preferred embodiment of the invention, the bond-reducing material is selectively deposited by a printing technique. Such printing techniques include, but are not limited to;
The mechanism of bond reduction, or controlled bonding, is dependant on the particular bonding method used. Such methods may include, but are not limited to, laser welding, diffusion bonding, surface modified chemical bonding, solvent assisted bonding, thermal laminating, chemical covalent or charged surface group bonding, mechanical interlocking, ultrasonic welding, die-electric bonding, microwave bonding, electrostatic or magnetic attraction, and adhesive bonding. In diffusion based bonding the printed layer acts as a full or partial barrier layer preventing the inter diffusion of molecules between the layers to be bonded. Similarly in chemical bonding or mechanical interlocking from localised melting, such as with solvent, laser, ultrasonic, die-electric, microwave, and laminating bonding methods, the printed layer may also act as barrier layer preventing portions of the two bonding surfaces from coming into contact. Alternatively the printed layer imparts a different chemical aspect to the proximal surfaces altering the bonding strength.
In a preferred embodiment of the invention at least one of the layers to be bonded comprises a polymer, such as a: polyolefin; Cyclo Olefin Polymer; polypropylene; polyethylene; low density polyethylene; high density polyethylene; polymethyl-methacrylate; polycarbonate; polyethylene terephthalate; polyethylene terephtalate glycol; polybutylene terephtalate; polystyrene; polyimide; polyetherimide; acrylonitrile butadiene styrene; polyurethane; polydimethylsiloxane; cellulose acetate; polyamide; polyether ether ketone; polyvinylchloride; polyvinylidene chloride; polyvinylidene fluoride; polymethylpentene; polysulfone; polytetrafluoroethylene; polyoxide methylene; nitrocellulose, nylons, acrylics, acetates, polyacrylamides, latex or silica particles, glass fibres resins or combinations thereof.
In one embodiment the printed bond-reducing layer is located on only one surface prior to bonding. For example in
In another embodiment of the invention the printed bond-reducing layer is on a nearby surface but not be in direct contact with the bonding area, and acts to reduce the bonding process in a region of the bonding surfaces. For example in laser welding the printed layer may act as a mask effectively shielding the region of interest, either partially or fully, from the laser beam.
In another embodiment of the invention the bond-reducing material may impart or change aspects of the electrostatic or magnetic characteristics of the surface to effect a change in bond strength.
In one embodiment the bond-reducing material changes the electrostatic properties of the surface to effect a change in bond strength.
Where r is the distance between the two charges and ke a proportionality constant, which is equal to approximately 9×109 Nm2/C2. A positive force implies a repulsive interaction, while a negative force implies an attractive interaction.
In another embodiment the bond-reducing material comprises magnetic, ferromagnetic or paramagnetic properties which can actively be used to effect a change in bond strength. An example is illustrated in
Where A is the area of each surface, in m2; H is their magnetizing field, in A/m; μ0 is the permeability of space, which equals 4π×10−7 T·m/A; and B is the flux density, in T.
The bond-reducing material can extend beyond the interface of the bonding surfaces. In another embodiment of the invention the printed deposition of the bond-reducing material extends beyond the bonded area between two surfaces to provide an interface coating between the coated surface and the microfluidic structure. This is particularly advantages where the adhesive layer would otherwise provide one of the microstructure surfaces and cause detrimental surface characteristics in microfluidic applications. Such advantages gained may include altering the surface toxicity, wettability, non-specific binding, topology, transparency or refractive index properties.
The invention is particularly advantages for area bonding methods such as diffusion, surface modified, solvent, and thermal laminating to avoid bonding of adjacent layers both inside and near the microstructures. Similarly the resolution of selective bonding techniques can be improved by using such printing layers. The table directly below describes the approximate resolutions and printed material thicknesses obtainable from current reel-to-reel printing methods.
0.8-2.5
indicates data missing or illegible when filed
In cases where the microstructures may be deformed during the manufacturing process, then the bond-reducing material can be used to prevent adhesion, and therefore prevent permanent deformation of the microstructures. For example, it is often problematic sealing a microchannel or chamber structure with a thin polymer layer (sheet, film or laminate) where the channel width is greater than the channels height without causing deformation during bonding process. Such problematic bonding processes include thermal diffusion and lamination through a roller nip. Where the channel widths and pressures are large enough then substantial deformation may occur and the opposing surfaces of the microchannel may come into contact. By coating the top and/or bottom of these structures (an example of which is shown in
In one embodiment the bond-reducing layer provides an outline of the microfluidic channel proximal to the bond edge.
In another embodiment a pressure relief or burst valve comprises a bond-reducing material.
a, 8b, and 8c illustrate a pressure relief or burst valve using one deformable layer.
In another embodiment of the invention a burst valve for storing and releasing reagents comprises a bond-reducing material. This effectively allows a seal to be formed in the microstructure at spatially predefined points, which may then remain sealed until an applied force is used to overcome the reduced bond strength at these predefined points. This invention enables an effective barrier layer to oxygen and water transmission until the burst valve is opened, which is critical for the long term storage and release of reagents.
In another embodiment a valve structure comprises a bond-reducing material.
An alternative embodiment of a check, or one-way, valve is shown in
In another aspect of the invention it is advantageous to form complex fluid handling systems containing pump and or valve components. In one embodiment of the invention a microfluidic pump structure comprises a bond-reducing material.
In another embodiment
In another aspect of the invention the bond-reducing material is used to prevent bonding to parts of integrated components within a microfluidic device. This is particularly important where the materials used for both the microfluidic device structure and the integrated components are compatible with the bonding process. For example
In another embodiment of the invention a microfluidic device comprises any combination of pump and or valve components, wherein a bond-reducing material is used.
The invention overcomes the limitations described in the application of affinity chromatography by providing a planar substrate with discrete optical detection flow cells that contain porous material and have connecting microchannels for fluid delivery and/or removal, and a method for making the same.
The invention uses a porous material inserted into a planar substrate where the flow to or from the porous media is enabled by at least one connecting microchannel, and where there is an optical detection means for measuring an analyte in the porous network.
There are numerous advantages of this invention. These include:
The invention enables simpler and lower cost manufacturing process to be employed where an otherwise smaller structure with smaller tolerances would be required to provide an equivalent microstructured flow cell. There are many commercially available colorimetric, absorption, fluorescence, and chemiluminescent chemicals available from many suppliers that may be used for optical detection in this invention. The porous optical flow cells can operate with a high sensitivity using either opaque or transparent porous materials. The optical light path through the porous network can be depicted by variations of the two cases shown in
The flow cells may be configured with optionally a source or detection optics, which can be located on opposing sides or the same side of the detection flow cell. No source system is required in the cases where chemiluminescence or other light generating assays are used. In one embodiment the source and detection systems are located on either side of the planar substrate. For example
In a preferred embodiment the porous material is arranged so that the flow through the porous structure is perpendicular to substrates surface.
In a preferred embodiment of the invention a single substrate may have multiple detection flow cells containing porous material. Instances of where assays require detection of multiple reagents include but are not limited to the reading of multiple samples, multiple analytes in the same sample, the use of control samples, calibration factors, and the assay replicates or repeating the same tests. The multiple detection flow cells 1801 may be arranged with interconnecting microchannels in either series 1802 or parallel 1803 configurations, or arranged with microchannels having flows independent to one another 1804, as depicted in
In a preferred aspect of the invention a device for improving optical detection is provided by incorporating a porous material inside a detection flow cell to increase the surface area for binding. An example of this is the use of a porous material as a solid phase for the binding of analytes in affinity chromatography. In one embodiment the porous material may be a polymer, glass or ceramic filter that may or may not be surface modified to provide controlled surface chemistries for binding. Such polymers may include, but are not limited to, Cyclo Olefin Polymer; polypropylene; polyethylene; low density polyethylene; high density polyethylene; polymethyl-methacrylate; polycarbonate; polyethylene terephthalate; polyethylene terephtalate glycol; polybutylene terephtalate; polystyrene; polyimide; polyetherimide; acrylonitrile butadiene styrene; polyurethane; polydimethylsiloxane; cellulose acetate; polyamide; polyether ether ketone; polyvinylchloride; polyvinylidene chloride; polyvinylidene fluoride; polymethylpentene; polysulfone; polytetrafluoroethylene; polyoxide methylene; nitrocellulose, nylons, acrylics, acetates, polyacrylamides, latex or silica particles, glass fibres or combinations thereof.
In one embodiment, alteration of the surface chemistry or the binding of surface coatings to the porous materials may be performed by a batch based process before they are inserted into the card. In an alternative embodiment, alteration of the surface chemistry or the binding of surface coatings to the porous materials may be performed after the card is manufactured by using a flow through protocol as described herein.
In one embodiment a surface activation step is used to activate the surface of the porous material. Surface activation involves altering the chemical groups present on the surface and the result is dependent on both the substrate and activation method. Examples of common chemical bond modifications include amine, carboxylic acid, and hydroxyl species. Industry standard methods for surface modification include corona discharge, wet chemical modification, plasmas using a variety of gases such as argon, oxygen, nitrogen, ethylene oxide, ammonia, acetone, methanol, and ethylenediamine.
In one embodiment the surface coating on the porous material is a multi-layer coating. The attachment of the layers can be covalent, electrostatic, or caused by physical entrapment and are well known to people skilled in the art of biochemistry and surface treatment. Examples of such layers include materials that contain an overall or localized charge (cationic or anionic), or are able to provide these charges when attached to a substrate or another coating, small molecules such as salts, biomolecules, neutral and charged polymers or polyelectrolytes, ligands, surfactants, and combinations thereof. Many types of polymers are often used to directly adhere to a surface.
In a one embodiment, one or more surface coating layers may include any of; surfactant, cationic surfactants, anionic surfactants, amphoteric surfactants, and fluorine containing surfactants, phosphate, polyethylenimine (PEI), poly(vinylimidazoline), quaternized polyacrylamide, polyvinylpyridine, poly(vinylpyrrolidone), polyvinylamines, polyallylamines, chitosan, polylysine, poly(acrylate trialkyl ammonia salt ester), cellulose, poly(acrylic acid) (PAA), polymethylacrylic acid, poly(styrenesulfonic acid), poly(vinylsulfonic acid), poly(toluene sulfonic acid), poly(methyl vinyl ether-alt-maleic acid), poly(glutamic acid), dextran sulfate, hyaluric acid, heparin, alginic acid, adipic acid, chemical dye, protein, enzyme, proteins, enzymes, lipids, hormones, peptides, nucleic acids, oligonucleic acids, DNA, RNA, sugars, and polysaccharides, immunoglobulins G (IgGs) and albumins, such as bovine serum albumin (BSA) and human serum albumin, peptide, isocyannated terminated polymers, including polyurethane, and poly(ethylene glycol) (PEG); epoxy-terminated polymers, including PEG and polysiloxanes; and hydroxylsuccimide terminated polymers. or a salt or ester thereof.
In one embodiment a layer of either Biotin or PEG can be coated to a porous material, where the porous material has a charged surface opposite to that of the functional group on the PEG or Biotin molecules. In another embodiment a layer of biomolecules such as proteins, enzymes, peptides, DNA, or RNA are electrostatically attached to the surface of the porous material.
The porous material is inserted into the substrate during the card assembly process. Adhesives or localised melting at the interface of the filter to the surrounding material may be used to affect a seal along the filter edges. In one preferred embodiment a pressure fit is used where the filter is compressed into a hole that is smaller in diameter than the filter. This compression fit requires no adhesive and ensures no fluid leaks around the edges of the filter material.
The substrate may be made of any suitable polymer such as: polyolefins; Cyclo Olefin Polymer; polypropylene; polyethylene; low density polyethylene; high density polyethylene; polymethyl-methacrylate; polycarbonate; polyethylene terephthalate; polyethylene terephtalate glycol; polybutylene terephtalate; polystyrene; polyimide; polyetherimide; acrylonitrile butadiene styrene; polyurethane; polydimethylsiloxane; cellulose acetate; polyamide; polyether ether ketone; polyvinylchloride; polyvinylidene chloride; polyvinylidene fluoride; polymethylpentene; polysulfone; polytetrafluoroethylene; polyoxide methylene; nitrocellulose, nylons, acrylics, acetates, polyacrylamides, latex or silica particles, glass fibres or combinations thereof.
The microfluidic channels may be formed in the substrate, or formed in an attached layer connected to the substrate
b) depicts a cross section of sealing layers 1905, 1906 and the substrate 1901 with the porous material 1904 prior to assembly. In some embodiments the substrate 1901 or sealing layers 1905, 1906 contain microchannels which may be formed by methods including, but not limited to, micromilling, laser based lithography and beam scanning, plasma etching, wet chemical UV lithography using photoresists, soft lithography, x-ray lithography, print-head deposition, soft lithography, stamping, embossing, compression molding, thermoforming, injection molding and reaction injection molding.
For many microfluidic applications it is advantageous to control the pumping, valving, or debubbling in these microfluidic devices. Examples of the constructions of components to perform these operations are described in WO 2007/060523, the entire contents of which are incorporated herein by reference. An example implementation is depicted in
In one embodiment of the invention a device comprises multiple inlet microchannels with on-way valves, and/or a debubbler, and/or at least one detection flow cell, wherein optionally microfluidic flow control is provided by independently controlled fluid lines external to the device. For example, the device of
The method of using flow control valves valve control to adjust the flow rate from a single source is particularly advantages for cost and size reduction of the external instrumentation by reducing the number of pump and valve components required for complex devices.
In one embodiment of the invention the device comprises multiple inlet microchannels with variable flow valves, and/or a debubbler, and/or at least one detection flow cell, wherein optionally a pressure source external to the device provides pressure driven flow which is varied by the flow control valves. For example
In one embodiment of the invention a device comprises onboard reagents, and/or microchannels with variable flow valves, and/or a debubbler, and/or at least one detection flow cell, wherein optionally a pressure source external to the device provides pressure driven flow which is varied by the flow control valves. For example,
In another example a microfluidic card comprises components for reagent storage, and/or mixing or rehydration, and/or a debubbler, and/or controlled dosing, and/or Flow control and passive valves, and/or at least one detection flow cell containing porous material. These fluid handling components can be reconfigured on different cards to provide for the needs of different assays. Such assays include, but are not limited to: immunoassays such as the indirect, sandwich, competitive, and reverse ELISA methods. In the example of
The reagent storage can be either in liquid or dried format. In one embodiment dried lyophilised reagents are placed into the reagent and reconstitution chambers and water is stored or added through the water chamber.
For shelf life and stability considerations it is often advantageous in diagnostic applications to provide the reagents in a dried format. Dried reagents and processes of lypholizing are commonly known to those skilled in the art. Methods of lypholization by cryogenic methods are described in U.S. Pat. Nos. 3,721,725, 3,932,943, 4,848,094, 4,655,047. Whilst U.S. Pat. Nos. 4,820,627, 4,678,812, 4,762,857 and 4,115,537 describe processes suitable for preparing particles for tableting into diagnostic reagents.
As an example of the preparation of a freeze dried sample a stock solution made to one litre with distilled water containing the following; 0.5 mg of detection antibody, 25.5 g Sodium Chloride as a stabiliser, 3 g Triton X-100 as a surfactant to control bubble formation during dissolution, 71.5 g HEPES as a zwitterionic organic chemical buffering agent, and 84 g polyethylene glycol (MW 20,000) to facilitate formation of the matrix structure during freeze drying and for the development of turbidity during analysis. The freezing process is well known to those skilled in the art, as an example the droplets are dispensed into a cryogenic liquid. The rehydration of these freeze dried reagent droplets can be achieved with approximately 10 microliters of a 14:1 ratio mixture of water and human serum.
In one embodiment of the invention a method for performing a an immunoassay comprising the steps of: a) cross-linking a primary antibody to functional group on the porous materials surface; f) blocking non-specific antibody binding sites; g) incubating a sample containing a protein specific for the primary antibody; h) adding a detection antibody; and i) detecting the detection antibody.
In one embodiment the surface activation and binding of the porous material may be performed in a protocol after the cartridge has been assembled. For example such a procedure may involve the following steps: i) flowing 100 μl of 100% ethanol at 25 μl/min; ii) flowing 100 μl of a 1:1 mix of ethanol and distilled water at 25 μl/min; iii) flowing 100 μl of distilled water at 25 μl/min; iv) flowing 100 μl of carbonate buffer (pH=9.5) at 25 μl/min; v) incubating 20 μl of capture antibody for 45 mins in the detection cell; vi) washing with 250 μl of blocking buffer (0.1% BSA) at 25 μl/min,
In one embodiment the surface activation and binding of the porous material may be performed in a batch based protocol before the cartridge has been assembled. For example such a procedure may involve performing the following steps with the filters completely immersed in a stirred solution at room temperature: i) 100% ethanol for 10 minutes; ii) 1:1 mix of ethanol and distilled water for 10 minutes; iii) distilled water for 10 minutes; iv) carbonate buffer (pH=9.5) for 10 minutes v) incubating 20 μg/ml of capture antibody per filter for 45 mins; vi) blocking buffer (0.1% BSA) for 10 minutes,
In one embodiment the protocol for detecting Botulism toxin involves preparing the porous material (sintered HDPE particles giving an average pore size of <20 micron and a porosity of approximately 40%) by one of the flow through or batch based methods described above. Then performing the following flow based assay through the detection flow cells and analysing the result either visually or with a simple photodiode and LED source detection system. The flow based detection assay involves i) washing with 50 μl carbonate buffer (pH=9.5) at 25 μl/min; ii) introducing 100 μl of sample at 25 μl/min; iii) addition of 100 μl Biotinylated anti-BotA at 25 μl/min; iv) addition of 100 μl Streptavidin-polyHRP at 25 μl/min; v) washing with 50 μl carbonate buffer (pH=9.5) at 25 μl/min; vi) addition of 100 μl TMBN substrate at 25 μl/min with concurrent detection.
In one embodiment a card is provided that contains reagent reservoirs and or interfaces to headers that contain fluid reservoirs. These reservoirs can be filled prior to or after insertion of the card into an instrument that contains the detection and or pneumatic interface. The example of
In one embodiment the microfluidic device has a size and detector flow cell layout compatible with standard microtiter plate based systems (ANSI/SBS 1-2004: Microplates; ANSI/SBS 2-2004: Microplates; ANSI/SBS 3-2004: Microplates; ANSI/SBS 4-2004: Microplates). For example
In an alternative embodiment the microfluidic device has a size and detection flow cell layout compatible with standard microtiter plate based systems and incorporates on-board reagents. For example
Throughout this specification (including any claims which follow), unless the context requires otherwise, the word ‘comprise’, and variations such as ‘comprises’ and ‘comprising’, will be understood to imply the inclusion of a stated integer or step or group of integers or steps but not the exclusion of any other integer or step or group of integers or steps.
The reference to any prior art in this specification is not, and should not be taken as, an acknowledgement or any form of suggestion that the prior art forms part of the common general knowledge.
| Number | Date | Country | Kind |
|---|---|---|---|
| 91247026 | Sep 2009 | US | national |
This application claims priority to U.S. provisional patent application No. 61/247,026, filed on 30 Sep. 2009, the entire contents of which are incorporated herein by reference.
| Filing Document | Filing Date | Country | Kind | 371c Date |
|---|---|---|---|---|
| PCT/AU2010/001283 | 9/30/2010 | WO | 00 | 3/30/2012 |
