Field of the invention

The invention relates to a microfluid sample delivery system comprising one or more channels passing fluid from an inlet, to a measuring site and further to an outlet, in particular the invention relates to a polymer-based microfluid sample delivery system operating without the use of pumps.

Background of the invention

In a wide varieties of applications it is desirable to perform measurements on a small fluid sample, such as a small liquid sample. Examples of such applications range from clinical testing or diagnosis e.g. in hospitals or specialised clinics to research activities in universities and industry laboratories, e.g. related to analysis of small amounts of chemical substances.

In fluid sample delivery systems, the sample portion may be transferred from a loading site to a measuring site and possible further to a discharge site. A sample portion may typically be macroscopic, e.g. a blood sample, whereas only a microscopic amount may be needed or desired at the measuring site in order to perform a measurement. A sub-sample portion may be loaded into a loading site, e.g. by means of a micropipette, from where it is transferred to the measuring site.

In order to transfer a liquid sample from a first site to a desired location various kinds of liquid handling systems have been proposed. For example, a microfluid transferring system comprising a micropump is disclosed in the published US application US2003/0215342.

The presence of a pump, however, complicates the fabrication of a device rendering such a device more expensive and possible complicated to use.

Summary of the invention

The present invention seeks to provide an improved microfluid sample delivery systems. It is an object of the present invention to provide a sample delivery system capable of delivery of different liquid samples to open measuring sites and thereby -facilitating multiple simultaneous detection e.g. in connection with screening of test samples.

It is another object of the present invention to provide a microfluid sample delivery system which may transfer fluid to a desired location without the use of pumps.

It is an even further object of the present invention to provide a sample cielivery system suitable to be loaded both by hand using standard equipment or suitable to be loaded by means of an automated loading system, e.g. in connection with high-throughput screening.

It is an even further object of the invention to provide an inexpensive device which is fast to fabricate.

It is an even further object of the invention to provide a sample delivery system which is simple to use, even for non-experts.

Accordingly there is provided, in a first aspect a sample delivery system comprising

one or more flow channels each having an inlet and an outlet, each inlet being connected to an open entrance reservoir and wherein the one or more flow channels are connected to a measuring site located between the inlet and the outlet so that sample fluid flowing from the inlet to the outlet of each channel passes the measuring site.

The sample delivery system is a system for transferring a fluid sample, s uch as a liquid sample from an entrance reservoir to a measuring site and further to an outlet for discharging or collecting the fluid sample. The measuring site is a site for attaching or connecting measuring equipment, such as a detector or a sensor. The measuring equipment may be any type of detector capable of detecting a desired property of the fluid sample, such as a physical, a chemical, a biological and/or a biochemical property. The geometric elaboration of the measuring site may be adapted to fit a specific type of device desirable to insert into the measuring site.

The measuring site may, however, also be a site for attaching or connecting equipment which is desirable to bring into contact or interaction with a fluid sample, e.g. equipment, such as part of equipment which is brought into contact with a liquid substance, e.g. for coating purposes or other purposes such as chemical altering of the surface of the equipment. A number of flow channels may be present, each of the channels having an inl et and an outlet, whereas the inlet is a separate inlet for each of the flow channels. The outlet may either be a separate outlet or a shared outlet. Separate outlets may be desired in connection with fluid substances which should not be mixed, whereas shared outlets may be desired for fluid substances that may be mixed.

The one or more inlets are connected to open entrance reservoirs, thus reservoirs in direct contact with the surroundings. The size of the opening of the entrance reservoirs may be large enough for easy loading by the human hand. By providing the remaining part of the system with dimensions suitable for handling minimal fluid quantities, an interface between the macroworld into the microrange has been provided. The entrance reservoir may however also be used in connection with automated loading, such as automated droplet injection from an inkjet-type device.

The sample fluid introduced at the entrance reservoir may flow from the inlet to the outlet by means of forces arising from surface tension between the fluid and an inner surface of the channel. Thus, the fluid may flow due to a capillary force or a capillary-like force. A driving force on a fluid in a channel may arise due to interaction between the fluid and an inner surface of the channel and/or due to a pressure difference across a surface of the fluid, the pressure difference resulting from a surface tension across the surface of the fluid. It may be a great advantage to drive fluid from a reservoir to a desired l ocation by means of forces arising from surface tension, since a pump may be avoided. The obtainable fluid flow may depend upon the geometry of the system and the material properties of the system. These parameters may be known and taken into account when designing a system for a specific purpose.

The system may further comprise one or more exit reservoirs for outlet connection. An exit reservoir may be provided in order to collect the sample fluid after passage of the measuring site. It may be an advantage to collect the sample fluid, e.g. for easy handling of a measurement, or in connection with toxic fluid samples. A single shared reservoir may be present or each of the flow channels may be connected to individual exit reservoirs.

The one or more channels may be connected to the same measuring site or m ay be connected to different measuring sites. Thus a sample delivery system with one or more measuring sites may be provided. The number of measuring sites may depend upon the intended type used for the sample delivery system. It is an advantage to provide different types of systems, both with respect to number of channels, number of exit reservoirs and number of measuring sites, since a more versatile system may be provided. A versatile system may be needed due to the large variety of applications of the present invention.

The material of the polymer-based sample delivery system may be any suitable type of natural or synthetic polymer-based material or co-polymer-based material. The polymer- based material may be a plastic material, such as a thermoplastic or a thermoset, or such as a so-called photoplastic, i.e. a plastic material that may be photolithographically processed. The material of polymer-based sample delivery system may be selected from the group consisting of: SU-8 based polymers, such as XP SU-8 polymer, polyimides or BCB cyclotene polymers and parylene, but many other polymer-based materials or plastic materials could be used. The chemical name of SU-8 is glycidyl ether of bisphenol A. SU-8 may be a suitable component for fabricating a sample delivery system since it has a high chemical resistance, capable of supporting very high aspect ratios and it is relatively easy and fast to process.

It may be an advantage to provide a sample delivery system in a polymer-based material since the fabrication process is rendered simple, cheap, fast and flexible. It is an advantage that a cheap device may be provided, e.g. since the system may be used to measure chemical and/or biological substances which may be difficult to, time consuming to or even toxic to clean off, rendering it desirable to provide a disposable device.

The polymer SU-8 and possible other polymers are hydrophobic and it may be an advantage to render parts of a system fabricated in a hydrophobic material hydrophilic.

An inner surface of one or more channels, possibly including the inner surface of one or more measuring sites, may be rendered hydrophilic. Only the inner surfaces of the one or more channels may be rendered hydrophilic, whereas the part of or the remaining part of the system may be (or remain) hydrophobic. In this way a leaking from one channel to another channel may be suppressed or avoided.

The hydrophilic rendering may be obtained by a liquid treatment of the inner surface of the one or more channels. The liquid treatment may be obtained by flushing the inner surface with liquid Ethanolamine or a liquid Ethanolamine solution, or any other suitable liquid. The inner surface of the one or more channels may be provided with a surface which with water forms a contact angle below 90°, such as below 70°, such as below 50°, such as below 30°, such as below 10°. The sample delivery system may be provided with a volume of the entrance reservoir below 100 μL, such as below 50 μL, such as below 30 μL, such as below 10 μL, such as below 500 nl_, such as below 100 nl_, such as below 500 fL, such as below 100 fL And wherein the cross-sectional area of each of the one or more flow channels may be below 10 mm2, such as below 1 mm2, such as below 500 μm2, such as below 100 μm2, such as below 1 μm2.

The dimensions of the entrance reservoir and the geometric aspects of the channels may be provided such that the fluid flow in the one or more channels are substantially laminar- The laminar fluid flow may be maintained as the fluid passes the measuring site. It may b>e an advantage to provide laminar flow of the sample liquid since a more controlled flow is obtained thereby. It may be a further advantage to provide laminar flow of the liquid as it passes the measuring site since different liquid may pass in close contact without being mixed.

The one or more channels may be open channels or the one or more channels may be closed channels. The channels may e.g. be provided with or without a lid.

The sample delivery system may further comprise a support on which the one or more flow channels and at least one entrance reservoir are supported. In this way a chip comprising the sample delivery system may be provided. The chip may comprise a number of sample delivery systems.

According to a second aspect of the invention, a method is provided where a sample fluid is provided from an inlet to an outlet, each inlet being connected to an open entrance reservoir and wherein the fluid flows in the one or more channels, the channels being connected to a measuring site located between the inlet and the outlet, the fluid passing the measuring site, and wherein the fluid flows from the open entrance reservoir to the outlet by means of forces arising from surface tension between the fluid and an inner surface of the channel.

The method may provide liquid flow in a sample delivery system according to the first aspect of the invention.

According to a third aspect of the invention is provided a method of fabricating a polymer- based sample delivery system comprising one or more flow channels each having an inlet and an outlet, each inlet being connected to an open entrance reservoir and wherein the one or more flow channels are connected to a measuring site located between the inlet and the outlet so that sample fluid flowing from the inlet to the outlet of each channel passes the measuring site, wherein the method being a photolithography process.

The method may provide a sample delivery system according to the first and/or second aspect of the invention.

It may be an advantage to use a material which can be structured by standard UV- lithography and is compatible with most clean room processes.

These and other aspects, features and/or advantages of the invention will be apparent from and elucidated with reference to the embodiments described hereinafter.

Brief description of the drawings

Preferred embodiments of the invention will now be described in details with reference to the drawings in which:

FIG. 1 shows two embodiments of sample delivery systems,

FIG. 2 shows a cross section of an entrance reservoir and part of a flow channel,

FIG. 3 illustrates selective coating of micro-cantilevers,

FIG. 4 shows a schematic illustration of a sensor device inserted into the measuring site in a measuring situation,

FIG. 5 illustrates process steps involved in a method of fabricating a SU-8 sample delivery system according to the present invention, and

FIG. 6 shows an electron microscopy photo of a closed channel.

Description of preferred embodiments

A schematic drawing of two embodiments of the present invention is provided in FIG. 1. FIG. IA illustrates a chip comprising a support 1 into which three entrance reservoirs 2 are provided. Each entrance reservoir is connected to a flow channel 3. The flow channel being intersected by a measuring site 4. The measuring site being a recess in the support adapted for receiving a sensor device. Each of the flow channels also being connected to exit reservoirs 5. The channels may be provided with a bending of 90° or any other angle. In such bendings of a channel the corners may be made smooth in order to avoid clogging from bubbles attaching to sharp corners.

In FIG. IB a different embodiment of the present invention is provided. Here the flow channels are intersected by a shared measuring site 6, and each of the channels discharges into a common exit reservoir 7. In large solid areas of SU-8, holes 10 may be provided. Such holes are stress relief holes provided in order to avoid the polymer to crack due to build up of stress. The size of the holes depend upon the design, a typical size may be such as 50 μm x 50 μm. Stress relief holes may also be provided in the system illustrated in FIG. IA, even though not explicitly illustrated.

The size of the chip may be a few centimetres along each side, the reservoirs may have a width of tens of millimetres, whereas the size of the measuring sites may be around a few hundred of micrometer along each side.

It is to be understood that the shape of the various elements as well as the number of the various elements of the sample delivery system is limited only by the accompanying claims. The fact that two specific embodiments are provided serves only for illustrative purposes.

In FIG. 2 a cross section of an entrance reservoir 20 and part of a flow channel 21 are illustrated. The cross section may e.g. be obtained along the line indicated by reference numeral 8 in FIG. IA. The reservoir and the channel are partly filled with a liquid 22. The proportions of the reservoir 20, the channel 21, the width of the support 23 and the width of the lid 24, are not drawn to scale and chosen only for illustrative purposes. Thus the reservoir may or may not be wider and/or deeper, the channel may or may not be broader and/or thinner, etc. The inner surface 25 of the flow channel is hydrophilic resulting in a curvature of the liquid column end as shown. Due to the surface tension between the inner surface of the flow channel and the liquid, a capillary force F arises and drives the liquid forward. Flow rates of a few mm/s may be obtained.

The sample delivery system may be used in connection with a sensor device. In FIG. 3 is an example provided where a sensor device is selectively coated using the sample delivery system of the present invention, and in FIG. 4 is an example provided where the sample delivery system is used in connection with a detection measurement using a sensor device. The system may be used in connection with micro-cantilever chips, however the system may be used in connection with other types of sensor devices, and as mentioned the measuring site may be formed in correspondence with the specific shape of the sensor device. In FIG. 3 a coating process is illustrated and the measuring sites may in principle be referred to as coating sites, however for consistency the measuring sites are referred to using the term "measuring site" throughout the text irrespective of the specific purpose.

In FIG. 3A a micro-cantilever chip 30 is illustrated. The outer boundary of the chip is marked by the reference numeral 31, and the chip comprises three groups 33-35 of cantilevers 32. The chip including the cantilevers may typically be fabricated in the polymer SU-8. The cantilevers are typically a few hundred μm long.

When using micro-cantilevers for biochemical detection, the cantilevers may be sensitised to allow for specific molecular binding during an experiment. This may be obtained by coating the cantilevers with a sensor molecular layer to which a single type, or a specific type, of molecules may bind. Such coating may be done by dipping all cantilevers in a solution containing the desired sensor molecules. However, in this way all cantilevers are coated with the same layer. Alternatively, each cantilever may be treated individually by using a micropipette. This is, however, a tedious and labour intensive process. The micro- cantilever as illustrated in FIG. 3A may in connection with the sample delivery system illustrated in FIG. IA be used to separately coat each group 33-35 of cantilevers in a fast way without any risk of cross-contamination.

In FIG. 3B the cantilever chip 30 is placed on top of the sample delivery system 36 in such a way that the groups of cantilevers are aligned with the corresponding measuring sites 38. Each measuring site being connected to a flow channel 37.

By tilting the cantilever chip 30 the groups of cantilevers are inserted into the cavity of the measuring sites, this is illustrated in FIG. 3C.

With the cantilevers inserted into the measuring sites, different solutions may be added into the entrance reservoirs e.g. by using a micropipette with a volume of up to 20 μL By capillary forces, the liquid is drawn into the flow channels and thereby the measuring sites.

As an example, DNA labelled with Cy3 was injected into the first reservoir and DNA labelled with Cy5 was injected into the middle reservoir. The cantilever chip was subsequently lowered into the measuring sites (as illustrated in FIG. 3C) and the DNA was adsorbed onto the cantilevers. In order to verify successful coating, the chip was fluorescence scanned with a Cy3 filter (FIG. 3D) and a Cy5 filter (FIG. 3E). In the fluorescence images it is seen that the cantilever group 300 was successfully coated with Cy3 labelled DNA, whereas the cantilever group 301 was successfully coated with Cy5 labelled DNA. FIG. 4 is a schematic illustration of a sensor device 40 inserted into the measuring site 41 in a measuring situation. Liquid (not shown) is flowing from the inlet flow channel 42 passing the measuring site 41 and being discharged via the outlet flow channel 43. The sensor device 40 is a cantilever based strain sensor fabricated in the polymer SU-8.

As seen from the side, the sensor device comprises a support structure 44 onto which a cantilever 45 is attached. A strain sensor element 46 is embedded into the cantilever beam. The sensor element is electrically connected to an external terminal 47. The external terminal may further be electrically connected to, or include, electronic equipment used in connection with the measurement.

In order to fully understand FIG. 4, the principle for molecular recognition of a cantilever is explained. One side of a cantilever is first covered by a sensor layer. In the figure the sensor layer is provided on the top side of the cantilever, however the layer may alternatively be provided on the bottom side. It is, however important that the two sides are provided with two different surfaces in order to induce a differential surface stress on the two sides upon molecular binding, since it is the differential surface stress that will cause the cantilever to bend. A sensor layer may provide selective adsorption, or immobilization of different molecular species.

In general molecular layers are known to induce surface stress when they bind to a surface, due to van der Waals, electrostatic or steric interactions. The difference in surface stress on opposite sides of the cantilever induces a bending of the cantilever, this bending may be measured, e.g. by use of a strain sensor, detecting the induced bending. Alternatively the bending may be detected by shining a laser onto the cantilever beam and detect the position of the reflected laser spot.

In connection with FIG. 4 static bending (static mode) of the cantilever is explained. However, the cantilever may also be operated in the dynamic mode. In dynamic mode detection, the cantilever is oscillated, e.g. by means of a piezo-oscillator attached to the support structure, the bending of the cantilever is monitored e.g. by measuring the oscillation frequency. From the measured signal, the changes in the resonant frequency of the cantilever from the immobilization of molecules onto the cantilever is deduced. The resonance frequency of a cantilever depends, inter alia, on the cantilever mass, near environment viscosity and surface stress.

In a more specific use of the strain sensor and the sample delivery system as illustrated in FIG. 4, a series of liquid samples may be monitored for the presence of a specific molecular species, for example in a screening measurement where liquid samples such as a blood samples may be monitored for the presence of a specific antibody, where the strain sensors are coated with a layer which provides selective bonding of this antibody. Blood samples from different persons are provided into different entrance reservoirs and the cantilever bending are monitored. In this way it may e.g. be determined that the centre blood sample contains the antibodies in question, whereas the other samples do not.

In another type of measurement a series of the same liquid sample is provided to all of the entrance reservoirs, whereas individual cantilevers, or possible individual groups of cantilevers (as described in connection with FIG. 3) have been coated with sensor layers which are sensible to different types of molecules. For example, in connection with the example given above concerning screening of blood samples for specific antibodies. Here the same blood sample may be screened for a variety of different antibodies, and the type of antibody found may be deduced from the possible bending of the cantilevers.

In FIG. 5 a method of fabricating an SU-8 sample delivery system according to the present invention is described. A silicon wafer 50, such as a 4" <100>-terminated Si wafer, is used during the process as a support of the sample delivery system. Firstly in FIG. 5A, a release layer 51 of Cr/Au/Cr is deposited onto the wafer. This metal layer will be used in a later process stage to release the system from the wafer in a wet chromium etch. In FIG. 5B a bottom layer 52 of SU-8 is spin-coated on top of the release layer, the thickness of the layers may be determined by the spinning parameters. The layer is pre-baked to evaporate most of the solvent. In FIG. 5C a centre layer 53 is provide by spinning a second layer on top of the bottom layer. In FIG. 5D the channels, reservoirs and measuring sites (seen schematically in cross-section 54) is patterned by UV exposure of the SU-8. The resist is post-baked to further promote the crosslinking of UV-exposed SU-8. Afterwards, the remaining non-crosslinked SU-8 is removed by immersion in an SU-8 developer (propylene glycol monomethyl ether acetate, PGMEA). On top of the centre layer a lid 55 is attached, e.g. by flipping a pre-fabricated lid onto the centre layer. The layer may be attached by gluing, or by pressing softly while heating the system. Alternatively, the sample delivery system may be provided without a lid so that open channels are provided. In FIG. 5F the sample delivery system is released in a wet chromium etch.

After the fabrication of the sample delivery system, the inner surfaces of the flow channels may be rendered hydrophilic. The rendering may be obtained by a liquid treatment where the channels are flushed with a suitable liquid.

In order to provide close channels but avoiding attaching a lid, it is possible to focus the UV-radiation inside an SU-8 layer and thereby pattern the channels below the surface. This may be obtained by only exposing the SU-8 structure to a low energy dose. Afterward the channels of non-crosslinked SU-8 may be flushed away by immersing the system in an SU- 8 developer. In FIG. 6 an electron microscopy photo of a closed channel 60 with side walls 61 and an overhanging wall or lid 62 is shown.

Although the present invention has been described in connection with preferred embodiments, it is not intended to be limited to the specific form set forth herein. Rather, the scope of the present invention is limited only by the accompanying claims.

In this section, certain specific details of the disclosed embodiment such as material choices, geometry or architecture of the device or parts of the device, techniques, measurement set-ups, etc., are set forth for purposes of explanation rather than limitation, so as to provide a clear and thorough understanding of the present invention. However, it should be understood readily by those skilled in this art, that the present invention may be practised in other embodiments which do not conform exactly to the details set forth herein, without departing significantly from the spirit and scope of this disclosure. Further, in this context, and for the purposes of brevity and clarity, detailed descriptions of well-known apparatus, circuits and methodology have been omitted so as to avoid unnecessary detail and possible confusion.

It will be appreciated that reference to the singular is also intended to encompass the plural and vice versa, and references to a specific numbers of features or devices are not to be construed as limiting the invention to that specific number of features or devices. Moreover, expressions such as "include", "comprise", "has", "have", "incorporate", "contain" and "encompass" are to be construed to be non-exclusive, namely such expressions are to be construed not to exclude other items being present.