Patent Application
20120190128
The present invention relates to a modular microfluidic sample preparation system where the system comprises:
Furthermore, the present invention relates to a method of mixing a sample fluid and an additive in a first preparation module and delivering said mixed sample fluid and additive to a second preparation module using a modular microfluidic sample preparation system.
Microfluidic systems are used in a number of laboratory automation applications, such as the preparation of fluid samples for further analysis. Such a microfluidic sample preparation system may be formed as a cartridge for insertion into a cooperating slot of an apparatus for performing the actual measurement and analysis of the sample prepared by the microfluidic system.
Furthermore, sample preparation of complex fluids, such as the preparation of blood samples for a specific measurement/analysis, often requires numerous steps. Some of these sample preparation steps may be of a general nature, and are usually performed in substantially the same manner for different types of measurements/analysis, whereas other sample preparation steps are specific to the specific measurement/analysis to be performed.
The numerous processing steps give rise to a complex microfluidic system that requires a large footprint for its implementation on a microfluidic chip. Such a complex microfluidic system is difficult to manufacture, and may lead to substantial problems in production, with low production yields as a consequence. Therefore, the manufacturing of such systems sets high standards for the capabilities of the manufacturing facilities.
In addition to that, different parts of the systems may require different capabilities of the manufacturing system, for example if active fluids need to be filled into reservoirs of a system, these typically need to be handled differently compared to less fragile parts of the system, e.g. common plastic parts. During the manufacturing of the parts for a microfluidic sample preparation system, the most demanding part sets the general standard of the parameters for all the parts of the system. Such parameters could e.g. be the level of cleanness in the production, sensitivity to heat, sensitivity to pressure or similar parameters. However, manufacturing the system by a standard, which is in fact too high for most of the parts, is expensive. Furthermore, the different parts of the system may require different storing facilities, e.g. if the system contains fluids that need storage in a controlled environment, e.g. with respect to moisture or temperature. Storing both sensitive parts and non-sensitive parts in a controlled environment takes up space in storage facilities and is inefficient as storing in a controlled environment is far more expensive than storing in a standard environment.
It is an object of the invention to provide a new and improved modular microfluidic sample preparation system which, at least partially, overcomes the disadvantages of the systems mentioned above.
This aspect is obtained by a modular microfluidic sample preparation system of the above mentioned type, wherein the first preparation module is connected to the second preparation module so that the second surface of the first preparation module is facing towards the first surface of the second preparation module and so that the outlet of the first preparation module faces the intake of the second preparation module.
Preparation steps performed by the modular microfluidic sample preparation system according to the invention may comprise dosing, mixing, adding additives or reagents, incubation, filtering, temperature stabilisation, presentation of the prepared sample at a measurement port, such as in a sample chamber with an optical window, and the like.
Hereby, the first preparation module and the second preparation module can be manufactured separately and be mutually connected afterwards. The preparation modules may demand a different level of e.g. cleanness in the manufacturing facilities. By separating the cartridge into at least two preparation modules, it is possible to manufacture the preparation modules at different locations, each complying with different production standard requirements. The outlet of the first preparation module is arranged such that a fluid e.g. a blood sample that has been preconditioned in the first module, is delivered from the outlet of the first preparation module and received by the intake of the second preparation module from where it is transported into the microfluidic channel system of the second preparation module.
Furthermore, by splitting the sample preparation system into preparation modules for different steps of the preparation process, the complexity of each of the separately produceable preparation modules is reduced, thereby reducing the complexity of the process for producing the preparation modules. This simplifies the production and thereby improves the production yield for the total sample preparation system.
The modular structure of the sample preparation system also allows for combining different types of first and second preparation modules. For example, the first preparation module may be designed for performing general steps such that the output from the first preparation module is a pre-configured fluid sample that is suited for use as input for different specific sample preparation steps. The specific sample preparation steps for different types of measurements/analyses may be implemented in the second preparation module. Alternatively, first preparation modules for different types of sample pre-configurations may be provided, which are compatible with the same type of second preparation module.
In another embodiment according to the invention, the first preparation module and the second preparation module may comprise connecting means for detachably connecting the first preparation module with the second preparation module. In this way, it is possible to use each of the modules together with other modular sample preparation systems. The first and the second preparation module being detachable, renders it possible to use the preparation modules together with other sample preparation modules or sample systems.
The preparation modules may be provided with releasable connection means, e.g. forming a snap lock connection. This allows for an easy assembly/configuration of the desired sample preparation system at the end user site by choosing and connecting the required types of first and second preparation modules. The above-mentioned easy combination/configuration of different types of preparation modules is thus also available to the end user.
Furthermore, in case of the malfunction of a preparation module, it is possible to exchange a preparation module with another. The connecting means furthermore provide that the preparation modules are correctly positioned in relation to each other. Furthermore, it is possible to store the preparation modules separately and connecting them just before use. In this way, it is possible to store the preparation modules in facilities just suitable for the specific preparation modules, thereby optimizing the storing costs. For example, different preparation modules may have different shelf lives depending on the content of reagents supplied in the module. If active or otherwise time sensitive content is comprised in a module, it is advantageous that such module be kept at a stock number just sufficient to supply the demand within the durability of such content. However, if a first preparation module compared to a second preparation module can be stored for a longer period, the manufacturing costs can be lowered by producing a higher number of less sensitive modules in one batch. Thus, the assembling i.e. connecting of the sensitive and the less sensitive preparation modules just before shipping, or even at the location of the end user may have an influence on the total costs of the modular microfluidic sample preparation system.
In one embodiment according to the invention, the first lateral faces of the first preparation module may comprise connecting means (e.g. a bead or a ridge) for connecting a recessed area of the second lateral faces of the second preparation module (or vice versa). In this way a simple and reliable system of connecting the two preparation modules is provided. The manufacturing of such connecting means can be carried out during a moulding process and is thus cheap to manufacture.
In a further embodiment of the invention, the preparation modules of the system, when connected, forms a cartridge. During use, the end user is faced with just one object to handle. Although the microfluidic sample preparation system is modular, the end user is still faced with just one object to handle.
Preferably, the cartridge comprises all reagents and/or additives to be mixed with the sample during the sample preparation process, thereby ensuring that only the fluid sample to be analysed needs to be handled and presented at the input port of the cartridge. The cartridge provides everything required during the sample preparation process and a measuring port for performing measurements on the prepared sample.
Preferably, such measurements are optical measurements, and the cartridge presents the prepared sample in a sample chamber/channel that is provided with an optical access port/window. The cartridge may be inserted into a cooperating analysis apparatus activating/driving the sample preparation process, e.g. by mechanical, electrical, and/or optical means, by radiation and/or temperature control. The analysing apparatus may further perform measurements on the prepared sample at the above-mentioned measurement port, such as optical or electrical measurements. By configuring the cartridge such as to integrate additive/reagent compartments, it is further achieved that any fluid handling may be confined to filling the cartridge with the sample to be analysed. Any fluid exchange between the cartridge and the analysis apparatus, or any fluid handling by the analysis apparatus may thus be avoided, thereby decreasing the complexity of the analysis apparatus, shrinking the lab-space footprint of the analysis apparatus, and reducing the need for cleaning and maintaining the analysis apparatus to a minimum.
In a further embodiment of the invention, the modular microfluidic sample preparation system may be a modular microfluidic blood sample preparation system.
In yet another embodiment according to the invention, the outlet of the first preparation module may be positioned at a distance from the intake of the second preparation module.
When connected, the first and second preparation modules are aligned such that the outlet of the first preparation module opens towards an intake opening of the second preparation module. The intake of the second preparation module comprises receiving means adapted to receive the fluid from the outlet of the first preparation module and delivery means to deliver the fluid to the microfluidic channel system of the second preparation module.
Typically, the area of the intake opening/receiving means of the intake of the second preparation (the second preparation module intake) is larger than the cross-sectional area of the outlet of the first preparation module (the first preparation module outlet). Providing a distance between the outlet and the surface of the receiving means of the second preparation module intake facilitates the lateral distribution of the fluid sample expelled from the first preparation module outlet over the reception means surface/intake opening of the second preparation module.
Fluid may be transferred from the first module to the second preparation module in the following manner; Fluid is expelled from the first preparation module outlet by a driving force, such as hydrostatic pressure applied in/to the first preparation module. The fluid expelled from the first preparation module outlet is accumulated around the outlet of the first preparation module before the fluid is getting into contact with the surface of the intake of the second preparation module. Upon contact with the surface of the second preparation module intake, the fluid may be distributed over the surface of the intake receiving means. Delivery means are provided in the intake for transferring the received fluid sample to the microfluidic channel system of the second preparation module. One advantage of the fluid transfer from the first preparation module to the second preparation module according to the present modular sample preparation system is that it does not require a sealed fluid interconnect between the microfluidic channel systems of the first and second preparation modules, or any critical pressure/fluid tight seal when attaching the first module to the second preparation module. Thereby it is achieved that the sample preparation system may easily be assembled from prefabricated first and second preparation modules at the end user site without requiring special production skills or dedicated equipment.
The receiving means of the second preparation module intake may be a sheet of woven/non-woven fibrous material adapted to at least partially absorb/imbibe/soak up the fluid sample. The sheet is arranged in the intake opening of the second preparation module and facing towards the outlet opening when the two preparation modules are connected. The sheet is dimensioned such that the fluid sample, when soaked up by the fibrous material, may reach an intake capillary connecting the receiving means to the microfluidic channel system of the second preparation module. The intake capillary may be arranged at one of the edges of the sheet of fibrous material. By capillary action, the intake capillary may retrieve the imbibed fluid sample—or in the case of a multiple component fluid at least one of the components of the multiple component fluid—and deliver it to the microfluidic channels system of the second preparation module for further processing.
The distance furthermore provides sufficient space if e.g. the intake of the second preparation module comprises swelling material that expands when getting into contact with a fluid. In this way, it is possible to use different materials at the intake of the second preparation module.
In an advantageous embodiment according to the invention, the distance between the outlet of the first preparation module to the intake of the second preparation module may be 0.05 mm-1 mm or 0.1 mm-0.9 mm or 0.15 mm-0.8 mm. In this way, a free space is created wherein fluid from the outlet of the first preparation module may bulge out, still being in contact with the outlet, before said fluid is delivered to the intake of the second preparation module. Furthermore, sufficient space for expansion is provided. If the intake of the second preparation module is e.g. a filter having a thickness of approximately 0.32 mm which expands due to swelling upon wetting, such filter may expand approximately 0.15 mm.
In another embodiment according to the invention, the intake of the second preparation module may comprise a blood separation filter. In this way, it is achieved that the intake of the second preparation module prepares the sample, e.g. blood, for the preparation of the sample following in the second preparation module. Thus, the intake of the second preparation module functions as a first step in the preparation carried out in the second preparation module.
Blood is a complex multiple component fluid. The separation of a multiple component fluid into different components may be performed in the intake of the second preparation module by providing receiving means and/or delivery means that are configured to preferably transport one or more desired components as compared to the remaining components present in the fluid sample received from the first preparation module. In particular, the blood filter may be configured to retain white and/or red blood cells while blood plasma is selectively transferred to the microfluidic channel system of the second preparation module.
The selective transport of the blood plasma in the filter may be provided by the capillary effect of the filter, preferably imbibing blood plasma while blood cells are retained by the crosslinked fibres of the filter material. An excess amount of mixed sample fluid, i.e. a blood sample pre-configured with the additive(s) of the first preparation module is provided to the intake of the second preparation module and on the filter, thereby saturating the filter material such that blood plasma reaches the intake capillary connected to the edge of the filter.
As mentioned above, a distance may be provided between the outlet of the first preparation module and the surface of the blood filter in the intake of the second preparation module. The distance allows for delivering the mixed fluid sample onto the blood filter without applying substantial pressure on the filter, thereby avoiding that blood cells are pressed through the filter into the second microfluidic channel system. The filter is dimensioned such that saturation of the filter material with blood plasma may be maintained with the amount of mixed sample fluid delivered to the intake of the second preparation module. Saturation should be maintained as long as needed to retrieve the desired amount of blood plasma into the second preparation module.
The capillary drag of the intake capillary in contact with the saturated filter material drags the fluid into the microfluidic channel system of the second preparation module. A capillary stop that may be provided in the second microfluidic channel system allows for limiting/controlling the volume of sample fluid, here blood plasma, transferred from the intake into the second module.
The blood filter may form the receiving means of the intake, wherein the blood filter is a sheet of filter web with a receiving portion arranged in the intake opening such that the receiving portion under operation faces the outlet opening. Typically, a transverse dimension of the receiving portion exceeds the corresponding transverse dimension of the outlet of the first preparation module. Advantageously, the filter web is therefore arranged at a distance from the outer edge of the outlet opening so as to improve the distribution of fluid over the receiving portion. Furthermore, the sheet of filter web may shaped so as to form a delivery tab, said delivery tab extending outwardly from the receiving portion and being connected to the intake capillary so as to establish a fluid communication between the filter web and the intake capillary.
According to one embodiment, the geometry of the blood filter may have a tapering outline, the wide portion of the tapering outline providing the receiving portion and the pointed portion forming the delivery tab. In this way, it is achieved that the blood filter is directing a fluid contained in the filter in a desired direction, e.g. towards the tapering part. In another embodiment, the filter may exhibit an efficiency of 5-50% or 10-25% output of plasma. The filter may e.g. have a thickness of 0.2 mm to 0.5 mm. Due to capillary drag in the filter, the filter may be wetted by the dispersion of fluid.
In yet another embodiment according to the invention, the outlet of the first preparation module may have a substantially circular cross section. In this way, the sample fluid delivered from the outlet of the first preparation module is equally distributed from the outlet when delivered from the outlet to the intake of the second preparation module. Due to the surface tension of the fluid, the bulge of fluid around the outlet of the first preparation module will be equally distributed around the outlet. The diameter of the outlet may be 0.5 mm-2 mm or 0.75 mm-1.75 mm or 1 mm-1.5 mm.
According to another embodiment of the invention, the outlet of the first preparation module may have a substantially oval cross section. In this way, it is achieved that the sample fluid delivered from the outlet of the first preparation module is delivered to the intake of the second preparation module so as to focus the flow in a certain direction.
In an additional embodiment according to the invention, the first preparation module may comprise a passive microfluidic mixing section in fluid communication with the first microfluidic channel system. In this way, it is achieved that fluids and/or additives supplied to the first preparation module can be mixed with the fluid provided through the inlet of the first preparation module.
In one embodiment according to the invention, the first preparation module may comprise an additive reservoir in fluid communication with the mixing section. In this way, it is possible for the first preparation module to comprise an additive to be added to the sample fluid. The additive reservoir being in fluid communication with the mixing section allows for the additive to be brought into the mixing section, e.g. by letting the sample fluid flow into the additive reservoir, thereby flushing the content of the additive reservoir out, or by forcing the content of the additive reservoir into the mixing section. Once the additive is brought into contact and mixed with the sample fluid, the mixture may require incubation in order to produce an appropriately mixed sample fluid that may be delivered to the second preparation module. For example, the additive may be a reagent comprising a marker, and an appropriate binding of the marker molecules to the target molecules may require such incubation during a given incubation time. Incubation may require temperature stabilisation. Required incubation times depend on the additive in question. For example, when analysing full blood as a sample fluid, the marker additive may be HRP requiring an incubation time in the order of a few minutes. Alternatively, the additive reservoir may comprise components suitable for providing molecular analysis of the sample by e.g. nucleic acid amplification. Such reagents may comprise e.g. nucleic acid polymerases, helicases, primers and/or probes. In one embodiment reagents suitable for analysing sample material through Polymerase Chain Reaction (PCR) are provided through the additive reservoir. In a more preferred embodiment, reagents suitable for analysing sample material through isothermal amplification are used. Such amplification techniques comprise e.g. Helicase Dependant Amplification (biohelix), Recombinase Polymerase Amplification (TwistDx), Nucleic acid sequence-based amplification (NASBA) (Mérieux), Stand displacement amplification (SDA; Becton Dickinson), Transcription mediated amplification (TMA; Gen-Probe), and Loop-mediated isothermal amplification (LAMP; Eiken).
In another embodiment according to the invention, the first preparation module may comprise an inlet reservoir in fluid communication with the inlet and the first microfluidic channel system of the first preparation module. In this way, it is possible to accumulate a sample fluid in the first preparation module.
In yet another embodiment according to the invention, the first preparation module may comprise a capillary stop delimiting the inlet reservoir from the mixing section. In this way it is achieved that the inlet reservoir of the first preparation module does not deliver its contents to the following mixing section unless a driving force, such as hydrostatic pressure, is applied forcing the content past the capillary stop. Thus, it is possible for the end user to fill sample fluid into the inlet reservoir without the sample fluid flowing into the mixing section. The inlet reservoir may thus be used for filling the sample preparation system with a pre-defined dosing volume of fluid. The pre-defined dosing volume may be determined by the total volume of the inlet reservoir. The total volume of the inlet reservoir may be less than 200 μl, alternatively less than 100 μl, and preferably about 50 μl.
Additionally according to the invention, the additive reservoir may comprise a plunger. In this way, it is achieved that the plunger seals the additive reservoir.
In another embodiment according to the invention, the plunger may be arranged such that the plunger can force the content of the additive reservoir into the mixing section. When forcing the plunger into the additive reservoir, the plunger will cause the content to be forced out of the additive reservoir. The plunger may cause a pressure to be built up in the additive reservoir. A wall, e.g. provided by a film or a foil, of the additive reservoir may, upon the build-up of a certain pressure, e.g. provided by pressing the plunger into the additive reservoir, allow the content of the additive reservoir to be delivered to the mixing section of the first preparation module.
In yet another embodiment according to the invention, the first preparation module may comprise closing means for shutting off the inlet of the first preparation module. In this way it is achieved that the sample fluid is kept in the first reservoir of the first module until a pumping pressure is applied. Thereby, the risk of contaminating other samples or operators is minimised. Furthermore, it is achieved that it is possible to let air, pass the closed inlet, e.g. for forcing the fluid around in the first preparation module, without the risk of air escaping through the inlet. In a further embodiment, the closing means may be a hinged lid. In this way, the closing means is kept near to the place of use and the risk of mislplacing the means is minimised. In another embodiment, the closing means may be an adhering foil or film. In yet another embodiment, the closing means may comprise a projection to be inserted in the inlet, thereby providing an air tight closing.
In a particular embodiment according to the invention, the first preparation module may comprise a transparent channel (visible to the end user). In this way, it is possible for the end user to easily determine, whether fluid enters the system, i.e. enters a part of the first microfluidic channel system.
In a further embodiment the transparent channel may be the inlet reservoir of the first preparation module. In this way, it is achieved that the user easily can determine whether the sample is sufficient in order to carry out an analysis of the sample fluid.
In an embodiment according to the invention at least one of the preparation modules may comprise identification means. In this way it is possible to adjust the machine analyzing the sample to the specific test to be performed. In a further embodiment according to the invention, the identification means may be a bar code, a chip or a RFID tag. In this way, the identification can carry a larger amount of information to be set in relation to the analysis to be performed, e.g. age of the preparation modules, type of content in reservoir(s) or batch number of content in reservoir(s).
Furthermore, in an embodiment according to the invention, the first preparation module may further comprise a pumping means, thereby ensuring that the fluid situated in the inlet reservoir of the first preparation module i.e. stopped by a capillary stop can be forced pass said capillary stop into the mixing section of the first preparation module. The pumping means may e.g. be an air filled bladder, a pump or be provided by a flexible membrane or foil covering an air filled reservoir or cavity of the first preparation module.
In one embodiment of the invention a capillary stop may delimit the inlet reservoir and the pumping means, thereby ensuring that the sample fluid does not enter the pumping means.
Furthermore, the invention relates to the use of a modular microfluidic sample preparation system according to any of above-mentioned embodiments for preparation of a blood sample. The sample prepared by using the microfluidic sample preparation system may subsequently be analysed in a blood analysing apparatus. To that purpose, the modular microfluidic sample preparation system may be inserted as a cartridge into a cooperating slot of such blood analysing apparatus. The blood analysing apparatus may further interact with the modular microfluidic sample preparation system so as to activate/drive the preparation process and analyse the sample. The interaction between the sample preparation system and the analysing apparatus may be e.g. mechanically, electrically, by radiation, heat and/or optically.
Furthermore, the invention relates to the use of a modular microfluidic sample preparation system in a blood analysing apparatus. The use of such microfluidic sample preparation system in a blood analysing apparatus provides that the end user only needs to fill-in a sample and the analysing apparatus will then perform the rest in order to carry out the analysis. The volume of the fluid provided to the modular microfluidic preparation system through the inlet port may be less than 100 μl.
Furthermore, the invention relates to a blood analysing apparatus using a modular microfluidic sample preparation system.
In another embodiment of a blood analysing apparatus according to the invention, the pumping means of a first preparation module may be activated by the blood analysing apparatus. In this way the specific time for performing the analysis is controlled by the analysing machine because the sample fluid is kept in the inlet reservoir, i.e. in a controlled non-damaging environment for the sample fluid, until the pumping means is activated.
Furthermore, the invention relates to a method of mixing a sample fluid and an additive in a first preparation module and delivering said mixed sample fluid and additive to a second preparation module using a modular microfluidic sample preparation system, the method comprising the steps of:
According to another method of preparing a blood sample using a modular microfluidic sample preparation system for analysing in a blood analysing apparatus, the method may comprise the steps of:
The invention has been described with reference to preferred embodiments. Many modifications are conceivable without thereby deviating from the scope of the invention. Modifications and variations obvious to those skilled in the art are considered to fall within the scope of the present invention.
