JP4852573B2 - Flow cell - Google Patents

Description

  The present invention relates to a flow cell including a measurement channel through which a chemical substance such as a biological substance, an environmental pollutant, and a health effect substance in a liquid to be measured is distributed and measured by a measurement device.

As a method for measuring a chemical substance in a sample liquid, for example, there is an optical measurement method in which a molecule-selective substance is immobilized in advance, and a sample liquid is allowed to flow there to selectively detect molecules to be bound. As a kind of optical measurement method, a method using a total reflection optical system is known. According to this method, excited evanescent waves are used as probe light, and coupling near the surface is directly and highly sensitive. Can be measured.
In such a total reflection optical system, surface plasmon resonance (SPR) utilizing the fact that an evanescent wave excited by total reflection resonates with and absorbs the surface plasmon on the surface of the metal thin film formed on the substrate surface. The measurement method is particularly well used.

  When measuring a sample liquid by the surface plasmon resonance measurement method, a fine channel is formed on a substrate, and probe molecules are immobilized on a metal film placed in the channel, Pass the sample liquid through. Then, based on the interaction between the probe molecule and the measurement target substance in the sample liquid at this time, it is measured whether or not the measurement target substance is contained in the sample liquid (see, for example, Patent Document 1).

  In such a measurement, since it is necessary to form the flow path on the substrate as described above and to provide other components on the same substrate, it is not easy to create. For example, when the sample liquid is transferred by pressure from the outside, parts such as a pump and a pipe are separately required in addition to the substrate constituting the flow path. As a result, waste of the sample liquid is generated due to the transfer path such as the piping, so that there is a limit to reducing the amount of the sample liquid.

In order to cope with this, there has been proposed a technique for forming a flow path for transferring a sample liquid or an area to be a pump by a capillary phenomenon between two opposing surfaces of two substrates by a microfabrication technique (for example, non-patent). Reference 1 or 2). A measurement chip (flow cell) created by this technique is provided with an inlet for introducing the sample liquid and a capillary pump for sucking the sample liquid. When the sample liquid is introduced into the inlet, The sample liquid sequentially flows out from the introduction port to the measurement channel and the pump, and when the sample liquid reaches the capillary pump, the sample liquid is sucked by a capillary phenomenon generated in the capillary pump. As a result, the sample liquid stored in the introduction port flows through the measurement flow path to the pump by the suction force of the capillary pump.
JP 2002-214131 A Martin Zimmermann, Heinz Schmid, Patrick Hunziker and Emmanuel Delamarche, “Capillary pumps for autonomous capillary systems”, The Royal Society of Chemistry 2007, Lab Chip, 2007, 7, 119-125.First published as an Advance Article on the web 17th October 2006 Anal.Chem. 1998, 70, 4974-4984

By the way, the measurement chip as described above is usually formed by engraving a flow path on one surface of one substrate and joining the other substrate to the one surface, and the sample liquid is blocked by two substrates. It is set as the structure distribute | circulated as the flow path in the remaining space.
However, in the case of such a configuration, there is a problem that if a gap is generated between the bonding surfaces of the two substrates, the sample liquid flows out to a region other than the flow path on the bonding surface due to the capillary force of the gap. Further, since the capillary force increases as the gap width becomes smaller, the sample liquid flows out of the flow path if only a slight gap is generated, as long as the close contact between the substrates cannot be made complete.
In addition, in order to eliminate the gap at the bonding surface, a method of forming one substrate with a soft material and bringing the substrates into close contact with each other by using elastic deformation of the substrate can be considered. Resulting in a decrease in productivity.

  The present invention has been made in view of such a problem, and it is possible to reliably prevent the sample liquid from entering a region other than the flow path and to allow the sample liquid to smoothly flow through the flow path. An object is to provide a simple flow cell.

In order to solve the above problems, the present invention proposes the following means.
That is, the flow cell according to the present invention includes a first substrate, a second substrate disposed on the first substrate, an inlet for introducing a sample liquid formed on the second substrate, and the first substrate. A flow cell that is formed between an upper surface of one substrate and a lower surface of the second substrate, has one end connected to the introduction port, and has a detection unit installed in the middle thereof; A microgap between a protrusion protruding from one surface of the upper surface of the first substrate and the lower surface of the second substrate toward the other surface and the other surface, and a capillary tube of the microgap The sample liquid is circulated so as not to flow out to a region outside the minute gap by force.

According to the flow cell having such a feature, the sample liquid introduced into the introduction port flows through the portion where the minute gap exists due to the capillary force of the minute gap between the protrusion on one surface and the other surface. Will circulate as.
In addition, by providing an abrupt (on the staircase) structural change between the minute gap and other regions, the contact angle of the liquid, which is uniquely determined by the characteristics of the channel material and the liquid, also changes abruptly. Is required. The sudden change in the contact angle acts as a resistance against the progress of the liquid, prevents the liquid from flowing out of the minute gap, and does not flow outside.

  The flow cell according to the present invention further includes a pump unit formed between the upper surface of the first substrate and the lower surface of the second substrate and connected to the other end of the flow path. Comprises a pump forming portion raised one step from either one of the upper surface of the first substrate and the lower surface of the second substrate toward the other surface, and between the pump forming portion and the other surface The sample liquid that has reached from the introduction port through the flow path is sucked by a capillary force due to the minute gap.

According to the flow cell having such a feature, when the sample liquid reaches the pump part, the sample liquid can be smoothly circulated in the flow path by the pump part being sucked by the capillary force.
Moreover, since the capillary force by the pump unit is generated by a minute gap between the pump forming unit and the opposing surface of the second substrate in the same manner as the above flow path, the sample liquid that reaches the pump unit is , It is possible to prevent outflow to an area other than the pump section.

  Furthermore, in the flow cell according to the present invention, the minute gap in the pump section may gradually increase as the distance from the connection portion with the flow path in the pump section increases in the depth direction.

  This makes it possible to hold more sample liquid and increase the capacity of the pump unit.

Furthermore, in the flow cell according to the present invention, a plurality of pillars that connect the pump forming portion and the other surface may be provided.
Since the surface area of the pump part is increased by the pillar and the capillary force is increased accordingly, the sample liquid in the channel can be sucked into the pump part more reliably and smoothly.

  According to the flow cell of the present invention, the sample liquid flow path is formed as a minute gap between the first substrate and the second substrate, and the sample liquid is circulated by the capillary force of the minute gap. It is possible to reliably prevent the sample liquid from flowing out to the region other than the flow path, and to smoothly circulate the sample liquid in the flow path.

  Hereinafter, a first embodiment of a flow cell according to the present invention will be described in detail with reference to FIGS. 1 to 6. 1 is a perspective view of the flow cell of the first embodiment, FIG. 2 is a plan view illustrating the internal structure of the flow cell of the first embodiment, FIG. 3 is an exploded perspective view of the flow cell of the first embodiment, and FIG. 2 is a cross-sectional view taken along line AA in FIG. 2, FIG. 5 is a cross-sectional view taken along line CC in FIG. 4, and FIG. 6 is a cross-sectional view taken along line BB in FIG.

  As shown in FIG. 1, the flow cell 1 of the present embodiment is arranged with a lower substrate (first substrate) 11 having a substantially plate shape having a rectangular shape in plan view, and an outer diameter of the lower substrate 11 being the same. The upper base (second substrate) 12 to be provided is formed by being laminated.

In such a flow cell 1, as shown in FIG. 2, the introduction part 13 into which the sample liquid S is introduced, and two pump parts 14 and 14 formed between the first substrate 11 and the second substrate 12 are provided. And the flow path 15 which connects the said pump part 14 and the introducing | transducing part 13 is provided.
Specifically, the flow path 15 is connected to the introduction section 13 at one end and irradiated with measurement light or the like from an external device, and is branched into two while being connected to the measurement flow path 16 at one end. The other end is composed of a connection channel 17 connected to the pump parts 14 and 14, respectively.

The first substrate 11 is formed from optical glass such as BK7, for example, and has a substantially rectangular shape in plan view with a plate thickness of about 1 mm and a side of about 16 mm. A metal thin film 20 is provided on the upper surface 11a of the first substrate 11 (the surface of the first substrate 11 facing the second substrate 12) 11a. For example, Au or the like is used as a material of the metal thin film 20, and the upper surface 11a of the first substrate 11 is formed by performing vapor deposition, sputtering, plating, or the like.
The metal thin film 20 may be formed only in a portion corresponding to the measurement flow channel 16.

The second substrate 12 is formed of a material such as acrylic having a thickness of about 0.5 to 5 mm, has a rectangular shape in plan view corresponding to the first substrate 11, and is closer to one side of the second substrate 12. An introduction port 12c is formed through the substantially central portion.
As shown in FIG. 3, the outer peripheral edge 12b of the lower surface 12a (surface facing the first substrate) 12a of this second substrate 12 is raised one step toward the first substrate 11, and the first substrate 11 and the second substrate When the 12 is joined, the outer peripheral edge portion 12 b comes into close contact with the upper surface 11 a of the first substrate 11. In the following description, the lower surface 12a of the second substrate 12 refers to a plane that is recessed by one step from the outer peripheral edge 12b, which is the inner side of the outer peripheral edge 12b.

  Further, an annular ridge 21 formed so that the lower surface 12a of the second substrate 12 protrudes one step toward the first substrate 11 is formed at the opening edge of the introduction port 12c on the lower surface 12a of the second substrate 12. Is formed. When the heights of the annular ridge 21 and the outer peripheral edge portion 12b based on the lower surface 12a are compared, the annular ridge 21 is slightly lower than the outer peripheral edge portion 12b. Is about 50 μm, for example.

The lower surface 12a of the second substrate 12 has one end connected to the annular ridge 21 and extending toward the other side opposite to the one side, and the linear ridge 22 A meandering ridge 23 meandering in the form of a crank while having one end connected to the other end and having a plurality of bent portions and extending to the vicinity of the other side, and a meandering ridge connected to the other end of the meandering ridge 23 Connection ridges 24 extending toward both sides of 23 are formed.
The straight ridges 22, the meandering ridges 23, and the connecting ridges 24 protrude to the first substrate 11 side up to the same height as the annular stepped portion 21 with the lower surface 12a as a reference.

  Furthermore, a pump forming portion 25 is formed on the lower surface 12 a of the second substrate 12 so as to be connected to each of the two connection protrusions 24. The pump forming portions 25 are raised one step up to the same height as the annular ridge 21 with respect to the lower surface 12a. Two pump forming portions 25 are arranged on the left and right objects with the linear ridge 22 as the axis of symmetry. It is formed in a rectangular shape in plan view with the extending direction of the strip 22 as the longitudinal direction. In addition, an air hole 26 penetrating the second substrate 12 in a substantially rectangular shape is formed on the one side of the pump forming portion 25.

  As described above, in the second substrate 12 of the present embodiment, the annular ridge 21, the linear ridge 22, the meandering ridge 23, the connection ridge 24, and the pump forming portion 25 are provided on the first substrate with respect to the lower surface 12a. The outer peripheral edge portion 12b is arranged so as to protrude further to the first substrate 11 side.

  Moreover, such a 2nd board | substrate 12 can be produced by the injection processing using the metal mold | die in which the predetermined pattern was formed, laser processing, cutting processing by an end mill, etc., for example.

The flow cell 1 in the present embodiment is formed by stacking the first substrate 11 and the second substrate 12 described above.
Specifically, the first substrate 11 is stacked from the upper surface 11a so that the lower surface 12a of the second substrate 12 faces the upper surface 11a, and the first substrate 11 is pressed from the upper surface side. At this time, an adhesive, an adhesive, a double-sided seal, or the like is interposed on the surface of the outer peripheral edge portion 12b of the second substrate 12, so that the upper surface 11a of the first substrate 11 and the outer peripheral edge portion 12b of the second substrate 12 are Are joined, and the first substrate 11 and the second substrate 12 are fixedly integrated.

At this time, in the second substrate 12, the annular ridge 21, the linear ridge 22, the meandering ridge 23, the connection ridge 24, and the pump forming portion 25 are slightly recessed from the outer peripheral edge portion 12b. A minute gap is formed between the annular step portion 21, the straight ridge 22, the meandering ridge 23, the connection ridge 24, the pump forming portion 25, and the upper surface 11 a of the first substrate 11. The distance of the minute gap is determined by the height difference between the annular step portion 21, the straight ridge 22, the meandering ridge 23, the connection ridge 24, and the outer peripheral edge portion 12b of the pump forming portion 25. Is about 50 μm.
In the flow cell 1 of the present embodiment, the minute gaps formed in this way are used as the introduction part 13, the measurement flow path 16, the connection flow path 17 and the pump part 14 shown in FIG. A flow cell 1 to be distributed is configured.

  In addition to the above, as a configuration for forming the minute gap, for example, the height of the outer peripheral edge portion 12b of the second substrate is formed to be substantially the same as that of the annular protrusion 21, the linear protrusion 22, and the like. The first substrate 11 and the second substrate 12 are joined with a double-sided seal or the like on the surface, and the annular step portion 21, the straight protrusion 22, etc. and the upper surface 11 a of the first substrate 11 are utilized by utilizing the thickness of this double-sided seal. A minute gap may be formed between the two. In this case, the distance of the minute gap can be easily adjusted by appropriately changing the thickness of the double-sided seal or the like.

Next, the operation of the flow cell 1 according to the present embodiment will be described.
When the sample liquid S is injected into the introduction port 12c, the sample liquid S enters the minute gap between the annular protrusion 21 of the second substrate 12 and the upper surface 11a of the first substrate 11, that is, the introduction portion 13 by capillary action. . Then, the sample liquid S progresses by the capillary action in the order of the measurement channel 16 and the connection channel 17 that are also minute gaps (see FIG. 4).

  At this time, the sample liquid S flows only through the measurement flow path 16 and the connection flow path 17 which are the regions where the micro gaps exist, and does not flow out to other portions. That is, for example, as shown in FIG. 5, there is an abrupt structural change between the straight ridge 16 that forms a minute gap and the lower surface 12a of the second substrate 12 that is recessed by one step from the straight ridge 16. When the liquid S tries to flow out of the minute gap, the contact angle between the sample liquid S and the second substrate 12 changes greatly. In this case, since the rapid change in the contact angle acts as a resistance against the progress of the liquid, the sample liquid S is retained in the minute gap and does not flow out to a region outside the minute gap. Therefore, the sample liquid S flows reliably and smoothly only in the measurement flow path 16 and the connection flow path 17 which are the formation regions of the minute gaps.

In this way, the sample liquid S that has passed through each flow path 15 flows into the pump units 14 and 14. Since the pump unit 14 is also formed by a minute gap, a capillary phenomenon appears, and the sample liquid S that reaches the pump unit 14 advances through the inside (see FIG. 6). Further, the sample liquid S is retained inside without flowing out of the formation region of the pump portion 14 by the same action as described above.
When the sample liquid S reaches the air hole 26, the contact angle between the sample liquid S and the pump forming portion 25 of the second substrate 12 changes greatly, and this acts as a resistance against the progress of the liquid. The progress of the liquid S is stopped. Thereby, the suction operation of the sample liquid S by the pump unit 14 ends.

In the flow cell 1 in which the sample liquid S is circulated in this way, the surface plasmon resonance phenomenon is caused in the measurement channel 16 in the process in which the sample liquid S is injected into the inlet 12c and sucked up to the pump units 14 and 14. Measurement of the sample liquid S used is performed.
The measurement using the surface plasmon resonance phenomenon uses the resonance of the evanescent wave and the surface plasmon wave on the surface of the metal (in this embodiment, the metal thin film 20) in contact with the specimen to be measured.

In this measurement, as shown in FIGS. 11 and 12, the flow cell 1 in which the light emitted from the light source 101 is collected by the incident side lens 102 and is incident on the prism 103 and is in close contact with the upper surface portion 104 of the prism 103. The metal thin film 20 that functions as a measurement unit is irradiated. The sample liquid S, which is a specimen, is arranged in contact with the surface of the metal thin film, and is condensed and irradiated with light transmitted through the first substrate 11 of the flow cell 1 on the back surface of the metal thin film 20.
The light irradiated in this manner is reflected at the interface between the first substrate 11 and the metal thin film 20, and the light intensity is measured by the light detection unit 106 made of an imaging element such as a so-called CCD image sensor, and the resonance occurs. A valley where the reflectance decreases with angle is observed.

  In such a measurement, the presence or absence of a sample that selectively binds to an antibody or DNA fragment fixed on the surface (detection unit side) of the metal thin film 20 is detected. A sample liquid is arranged in the detection unit. In the state, there is no distinction between a change due to the reaction between the target sample and the antibody and a change due to a state in which foreign matter has settled and accumulated on the detection unit. On the other hand, by allowing the sample liquid to flow on the metal thin film 20, the sedimentation of the foreign matter is suppressed, and the change due to the above-described reaction is selectively detected to reliably perform the measurement. Can be done.

  As described above, in the flow cell 1 of the present embodiment, the sample liquid S is formed by the capillary force of these minute gaps using the minute gap between the first substrate 11 and the second substrate 12 as the flow path 15 and the pump unit 14. Thus, it is possible to reliably prevent the sample liquid S from flowing out to the region other than the flow path 15 and the pump unit 14.

Also, for example, when a flow cell is formed by engraving a flow path on one surface of one substrate and bonding the other substrate to the one surface, if a gap occurs on the bonding surface, the sample liquid S is generated by the capillary force of the gap. Will invade areas other than the flow path on the joint surface. Therefore, in such a case, since it is necessary to perform close contact between both substrates, the production process becomes technically complicated, and it is necessary to select a material capable of close contact, which increases production costs. .
However, in the flow cell 1 of the present embodiment, since the gap is intentionally formed and used as the flow path 15, the adhesion between the contact surfaces of both the substrates 11 and 12 is not reliable, and there are some gaps on the contact surfaces. Even if it occurs, the flow path 15 and the pump unit 14 are not affected. Therefore, it is possible to facilitate production and reduce production costs.

As a modification of the first embodiment, the pump forming part 25 in the second substrate 12 is connected to the air hole 26 from the connection part with the flow path 15 in the pump part 14 as shown in FIG. It may be inclined in a direction away from the first substrate 11 as it goes.
As a result, since the minute gap becomes larger toward the back of the pump unit 14, more sample liquid S can be retained in the pump unit 14, and the capacity of the pump unit 14 can be increased. .

  Next, a flow cell 30 according to a second embodiment of the present invention will be described with reference to FIGS. FIG. 8 is a plan view showing the internal structure of the flow cell according to the second embodiment, and FIG. 9 is a sectional view taken along the line DD in FIG. In FIG.8 and FIG.9, the same code | symbol is attached | subjected to the same structural member as the flow cell 1 of 1st Embodiment, and detailed description is abbreviate | omitted.

The flow cell 30 of the present embodiment is different from the pump unit 14 of the first embodiment with respect to the configuration of the pump unit 31, and the pump unit 31 includes the first substrate 11 and the first substrate 11 as shown in FIGS. A plurality of pillars 32 that connect the two substrates 12 are provided.
That is, in the present embodiment, a plurality of substantially cylindrical pillars 32 that connect the pump forming portion 25 of the second substrate 12 and the upper surface 11a of the first substrate 11 in the thickness direction of the flow cell 30 are arranged at equal intervals. ing. As a result, the surface area per unit volume of the pump unit 31 is increased as compared with the case where the pillar 32 is not provided. Therefore, a larger capillary force acts on the sample liquid S passing between the pillars 32, and the sample liquid S can be sucked more smoothly and reliably.

Further, as a modification of the flow cell 30 of the second embodiment, as in the first embodiment, for example, as shown in FIG. 10, the pump forming unit 25 is connected to the flow path 15 in the pump unit 31. It may be inclined in a direction away from the first substrate 11 toward the air hole 26. Thereby, the capacity of the sample liquid S of the pump unit 31 can be increased.
The shape of the pillar 32 is not limited to a cylindrical shape, and may be a polygonal shape or other shapes.
Furthermore, instead of the pillar 32 or together with the pillar 32, a plurality of protrusions for connecting the first substrate 11 and the second substrate 12 are provided, and a plurality of thin flow paths are formed in the pump portion 31 by the protrusions. An integrated structure may be formed.

As mentioned above, although the flow cells 1 and 30 which concern on embodiment of this invention were explained in full detail with reference to drawings, a concrete structure is not restricted to this embodiment, The design change of the range which does not deviate from the meaning of this invention Is also included.
For example, in the present embodiment, two pump parts 14 and 31 are formed. However, the present invention is not limited to this, and a single or three or more pump parts 14 and 31 are provided. There may be. Further, the flow path 15 is not limited to the aspect of the present embodiment, and the shape can be appropriately changed according to the design.
That is, in the flow cells 1 and 30 according to the present embodiment, it is essential to use a minute gap formed intentionally between the two substrates 11 and 12 as the flow path 15, and such technical contents are embodied. Thus, any embodiment is encompassed.

  In the embodiment, the measurement using the surface plasmon resonance reduction has been described. However, the present invention is not limited to this, and any other optical measurement method can be used as long as it can handle the sample liquid S. Can also be applied.

The perspective view of the flow cell of a 1st embodiment The top view explaining the internal structure of the flow cell of 1st Embodiment The disassembled perspective view of the flow cell of 1st Embodiment It is AA sectional drawing of FIG. It is CC sectional drawing of FIG. It is BB sectional drawing of FIG. It is a figure explaining the modification of 1st Embodiment. It is a top view explaining the internal structure of 2nd Embodiment. It is DD sectional drawing of FIG. It is a figure explaining the modification of 2nd Embodiment. It is a block diagram which shows the structural example of an SPR measuring apparatus. It is an enlarged view of the flow cell in FIG.

Explanation of symbols

DESCRIPTION OF SYMBOLS 1 Flow cell 11 1st board | substrate 12 2nd board | substrate 12c Inlet 15 Flow path 16 Measurement flow path 17 Connection flow path 14 Pump part 22 Straight protrusion (protrusion)
23 Meandering ridges (ridges)
24 Connecting ridges (ridges)
25 Pump formation part 20 Metal thin film (detection part)
30 Flow cell 31 Pump part 32 Pillar 33 Pump formation part

Claims (4)

A first substrate;
A second substrate disposed on the first substrate;
An inlet for introducing a sample liquid formed on the second substrate;
In a flow cell comprising a flow path formed between an upper surface of the first substrate and a lower surface of the second substrate, one end side of which is connected to the introduction port, and a detection unit disposed in the middle position,
The flow path is a minute gap between a ridge protruding from one surface of the upper surface of the first substrate and the lower surface of the second substrate toward the other surface and the other surface. The flow cell is characterized in that the sample liquid is circulated so as not to flow out to a region outside the minute gap due to the capillary force of the minute gap . A pump unit formed between the upper surface of the first substrate and the lower surface of the second substrate and connected to the other end of the flow path;
The pump portion includes a pump forming portion that is raised by one step from one surface of the upper surface of the first substrate and the lower surface of the second substrate toward the other surface,
2. The flow cell according to claim 1, wherein the sample liquid that has reached through the flow path from the introduction port is sucked by a capillary force caused by a minute gap between the pump forming portion and the other surface.   The flow cell according to claim 2, wherein the minute gap in the pump part gradually increases in a direction away from a connection point with the flow path in the pump part.   The flow cell according to claim 2 or 3, wherein a plurality of pillars for connecting the pump forming part and the other surface are provided.

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