OBSERVATION CHIP

An observation chip 1 includes: a plurality of substrates 10 stacked in a thickness direction Z of the substrates; a first channel 25 extending along a first axis O1 along the substrates; a surrounding channel 30 surrounding the first channel all around in a cross section orthogonal to the first axis in the first channel from an end of the first channel to an overall merging point P2 of the first channel, integrated with the first channel, and forming a second channel 55 that extends along a second axis O2 along the substrates; and an observation section 60 provided on a side farther away from the first channel than the overall merging point in the second channel, and transmitting electromagnetic waves from outside to the second channel. A length of the second channel in the thickness direction is constant from the overall merging point to the observation section.

The present application is based on, and claims priority from JP Application Serial Number 2023-039902, filed Mar. 14, 2023, the disclosure of which is hereby incorporated by reference herein in its entirety.

BACKGROUND

1. Technical Field

The present disclosure relates to an observation chip.

2. Related Art

Flow cytometers are used to detect, count, and sort cells, particles, and the like. In a flow cytometer, a sample liquid (sample fluid) containing cells is introduced into an observation chip (sorting device) that has fine channels formed inside. Then, the characteristics of each cell are analyzed using laser light or the like, and the cells are sorted according to the analysis results.

A known configuration of the observation chip is made by sealing the upper and lower sides of a silicon substrate, which has a plurality of channels, with glass substrates, as described in Non-Patent Document 1 (The Royal Society of Chemistry 2017, “On-chip cell sorting by high-speed local-flow control using dual membrane pumps”, Lab Chip, 2017, 17, 2760-2767), which is an example of the related art. In this observation chip, a channel for introducing a sheath liquid into a sample channel from above and below (vertical direction) and a channel for introducing a sheath liquid from left and right (horizontal direction) are formed.

However, when observing the sample fluid in the observation chip of Non-Patent Document1, the flow of the sample fluid flowing through the observation chip may be disturbed, and the sample fluid may not be observed properly.

SUMMARY

In order to solve the above problems, the present disclosure proposes the following means.

(1) The first aspect of the present disclosure provides an observation chip, including: a plurality of substrates stacked in a thickness direction of the plurality of substrates; a first channel extending along a first axis along the plurality of substrates; a surrounding channel surrounding the first channel all around in a cross section orthogonal to the first axis in the first channel from an end of the first channel to an overall merging point of the first channel, integrated with the first channel, and forming a second channel that extends along a second axis along the plurality of substrates; and an observation section provided on a side farther away from the first channel than the overall merging point in the second channel, and transmitting electromagnetic waves from outside to the second channel. A length of the second channel in the thickness direction is constant from the overall merging point to the observation section.

DESCRIPTION OF EXEMPLARY EMBODIMENTS

Hereinafter, an embodiment of the observation chip according to the present disclosure will be described with reference toFIG.1toFIG.12.

As shown inFIG.1andFIG.2, an observation chip1of this embodiment includes a plurality of substrates10stacked in a thickness direction Z of the plurality of substrates10.

As shown inFIG.2, for example, the plurality of substrates10include a channel substrate11, a channel substrate12, a channel substrate13, a protective substrate16, and a protective substrate17. That is, the plurality of substrates10include three channel substrates11,12, and13. For example, the channel substrates11,12, and13are formed in a rectangular shape that is long in a predetermined direction when viewed in the thickness direction Z. The channel substrates11,12, and13are made of metal such as stainless steel. The channel substrates11,12, and13may be made of resin having a Young's modulus of about 2 GPa (gigapascals), metal having a Young's modulus of about 70 GPa to 500 GPa, glass, ceramics, or the like.

For example, the thickness of the channel substrates11,12, and13is 0.1 mm or less.

Hereinafter, the side of the channel substrate11with respect to the channel substrate12is referred to as a first side Z1in the thickness direction Z (also simply referred to as the first side Z1). The side of the channel substrate13with respect to the channel substrate12is referred to as a second side Z2in the thickness direction Z (also simply referred to as the second side Z2). That is, the channel substrates11,12, and13are arranged in the order of the channel substrates11,12, and13from the first side Z1toward the second side Z2. It should be noted that there is no limit to the number of channel substrates included in the observation chip1, which may be one or two, or four or more.

The protective substrates16and17are made of transparent glass or the like and are formed to have the same external shape as the channel substrates11,12, and13. The protective substrate16is arranged closer to the first side Z1than the channel substrate11. The protective substrate17is arranged closer to the second side Z2than the channel substrate13.

That is, the protective substrate16, the channel substrate11, the channel substrate12, the channel substrate13, and the protective substrate17are arranged in this order from the first side Z1toward the second side Z2. The substrates10adjacent to each other in the thickness direction Z are bonded to each other by a known adhesive (not shown).

The channel substrates11,12, and13are each formed with a slit (reference numeral omitted) that penetrates in the thickness direction Z. By stacking the protective substrates16and17on the channel substrates11,12, and13that have slits, etc., as shown inFIG.1andFIG.2, the plurality of substrates10include a sample fluid supply section20, a first channel25, a surrounding channel30, a second channel55, an observation section60, a third channel65, and a separation section70.

As shown inFIG.1, the sample fluid supply section20is formed in a columnar shape. For example, the sample fluid supply section20is formed in the channel substrate12. An opening21communicates with the sample fluid supply section20. For example, the opening21is formed in the channel substrate13and the protective substrate17.

Here, as shown inFIG.1andFIG.2, a first axis O1is defined along the plurality of substrates10and along a predetermined direction. A reference plane S1that includes the first axis O1and extends along the thickness direction Z is defined. A cross section S2orthogonal to the first axis O1is defined (seeFIG.2). A direction orthogonal to the first axis O1and the thickness direction Z respectively is defined as an orthogonal direction (first orthogonal direction) Y.

The first channel25extends from the sample fluid supply section20along the first axis O1. As shown inFIG.3, the cross-sectional shape of the first channel25along the cross section S2(hereinafter referred to as a first channel cross-sectional shape) is rectangular. The cross-sectional shapes of the first channel25, etc. inFIG.3and subsequent drawings show a schematic configuration. For example, the first channel25is formed in the channel substrate12.

As shown inFIG.1andFIG.2, the side of the sample fluid supply section20with respect to the first channel25along the first axis O1is defined as an upstream side X1. The side of the first channel25with respect to the sample fluid supply section20along the first axis O1is defined as a downstream side X2. The sample fluid supply section20is provided at an end of the first channel25on the upstream side X1.

As shown inFIG.1andFIG.2, the surrounding channel30includes a pair of first branch channels31A and31B and a pair of second branch channels41and46.

In this embodiment, the configuration of the first branch channel31A and the configuration of the first branch channel31B are plane symmetrical with respect to the first reference plane S1. Therefore, the configuration of the first branch channel31A is indicated by adding a capital letter “A” to the number of the reference numeral. The configuration of the first branch channel31B, which corresponds to the first branch channel31A, is indicated by adding a capital letter “B” to the same number as the first branch channel31A. Thus, redundant description will be omitted. For example, a straight portion32A of the first branch channel31A, which will be described later, and a straight portion32B of the first branch channel31B are plane symmetrical with respect to the first reference plane S1.

The first branch channel31A includes the straight portion32A, a bent portion33A, and a bent portion34A.

The straight portion32A extends along the first axis O1. The straight portion32A is arranged at a position spaced apart from the first channel25on a first side Y1in an orthogonal direction Y orthogonal to the first channel25(hereinafter also simply referred to as the first side Y1).

The bent portion33A protrudes from an end of the straight portion32A on the downstream side X2toward a second side Y2opposite to the first side Y1in the orthogonal direction Y (hereinafter also simply referred to as the second side Y2). An end of the bent portion33A on the second side Y2is continuous from the first side Y1of the first channel25to the first channel25at a partial merging point P1in the first channel25.

As shown inFIG.1, the bent portion34A protrudes from an end of the straight portion32A on the upstream side X1toward the second side Y2and onto the first axis O1.

The first branch channel31B includes the straight portion32B, a bent portion33B, and a bent portion34B, which are configured in the same manner as the straight portion32A, the bent portion33A, and the bent portion34A of the first branch channel31A.

The straight portion32B is arranged at a position spaced apart from the first channel25on the second side Y2with respect to the first channel25. An end of the bent portion33B on the first side Y1is continuous from the second side Y2of the first channel25to the first channel25at the partial merging point P1in the first channel25.

The bent portion34B communicates with the bent portion34A.

As described above, the first branch channels31A and31B sandwich the first channel25in the orthogonal direction Y at the partial merging point PI of the first channel25.

As shown inFIG.4, the cross-sectional shape of the first channel25and the first branch channels31A and31B along the cross section S2at the partial merging point P1(hereinafter referred to as a partial merging point cross-sectional shape) is rectangular.

A length L13of the partial merging point cross-sectional shape in the thickness direction Z is equal to a length L11of the first channel cross-sectional shape in the thickness direction Z (seeFIG.3). A length L14of the first channel25in the orthogonal direction Y at the partial merging point P1is substantially the same as a length L12of the first channel cross-sectional shape in the orthogonal direction Y (seeFIG.3). A length L15of the partial merging point cross-sectional shape in the orthogonal direction Y is longer than a length L12of the first channel cross-sectional shape in the orthogonal direction Y.

For example, the first branch channels31A and31B are formed in the channel substrate12.

As shown inFIG.1, the first branch channels31A and31B are plane symmetrical with respect to the first reference plane S1.

For example, an opening36communicates with the portions of the bent portions34A and34B that communicate with each other. For example, the opening36is formed in the channel substrate13and the protective substrate17.

For example, the first branch channels31A and31B are formed in the channel substrate12.

As shown inFIG.1andFIG.2, the second branch channel41includes a straight portion42, a bent portion43, and a bent portion44.

The straight portion42extends along the first axis O1. The straight portion42is arranged at a position spaced apart from the straight portion32A on the first side Y1with respect to the straight portion32A of the first branch channel31A.

The bent portion43protrudes from an end of the straight portion42on the downstream side X2toward the direction between the second side Y2and the upstream side X1. An end of the bent portion43on the second side Y2is continuous from the second side Z2of the first channel25to the first channel25at an overall merging point P2in the first channel25. The overall merging point P2is located on the downstream side X2(on the side of the second channel55) of the partial merging point P1in the first channel25. In other words, the partial merging point P1is located on the side farther away from the second channel55than the overall merging point P2in the first channel25.

As shown inFIG.1, the bent portion44protrudes from an end of the straight portion42on the upstream side X1toward the second side Y2and onto the first axis O1. The bent portion44is arranged closer to the first side Y1than the bent portion34A of the first branch channel31A.

For example, the straight portion42and the bent portion43are formed in the channel substrate13, and the bent portion44is formed in the channel substrate12.

The second branch channel46includes a straight portion47, a bent portion48, and a bent portion49, which are configured in the same manner as the straight portion42, the bent portion43, and the bent portion44of the second branch channel41.

The straight portion47extends along the first axis O1. The straight portion47is arranged at a position spaced apart from the straight portion32B on the second side Y2with respect to the straight portion32B.

The bent portion48protrudes from an end of the straight portion47on the downstream side X2toward the direction between the first side Y1and the upstream side X1. An end of the bent portion48on the first side Y1is continuous from the first side Z1of the first channel25to the first channel25at the overall merging point P2in the first channel25.

The bent portion49protrudes from an end of the straight portion47on the upstream side X1toward the first side Y1and communicates with the bent portion44.

The second branch channels41and46may be plane symmetrical with respect to the reference plane S1.

As described above, the second branch channels41and46sandwich the first channel25in the thickness direction Z at the overall merging point P2of the first channel25. The first branch channels31A and31B and the second branch channels41and46of the surrounding channel30surround the first channel25all around in the cross section S2of the first channel25from the end of the first channel25on the upstream side X1to the overall merging point P2of the first channel25. Then, the first branch channels31A and31B and the second branch channels41and46are integrated with the first channel25and form the second channel55that extends along a second axis O2along the plurality of substrates10.

In this embodiment, the second axis O2coincides with the first axis O1. It should be noted that the second axis O2may intersect the first axis O1on a plane along the plurality of substrates10.

In this embodiment, a second orthogonal direction that is orthogonal to the second axis O2and the thickness direction Z respectively is parallel to the orthogonal direction Y. It should be noted that the second orthogonal direction may intersect the orthogonal direction Y.

As shown inFIG.5, the cross-sectional shape of the first channel25and the first branch channels31A and31B along the cross section S2at an intermediate portion between the partial merging point P1and the overall merging point P2(hereinafter referred to as a first channel intermediate portion cross-sectional shape) is rectangular. A length L17of the first channel intermediate portion cross-sectional shape in the thickness direction Z is equal to the length L13of the partial merging point cross-sectional shape in the thickness direction Z. A length L18of the first channel intermediate portion cross-sectional shape in the orthogonal direction Y is shorter than the length L15of the partial merging point cross-sectional shape in the orthogonal direction Y, and is substantially the same as the length L12of the first channel cross-sectional shape in the orthogonal direction Y. That is, the length L18of the first channel25and the first branch channels31A and31B in the orthogonal direction Y at the intermediate portion between the partial merging point P1and the overall merging point P2is shorter than the length L15of the first channel25and the first branch channels31A and31B in the orthogonal direction Y at the partial merging point P1.

It should be noted that the length L18of the first channel25and the first branch channels31A and31B in the orthogonal direction Y at the intermediate portion between the partial merging point P1and the overall merging point P2may be equal to the length L15of the first channel25and the first branch channels31A and31B in the orthogonal direction Y at the partial merging point P1.

As shown inFIG.6, the cross-sectional shape at the overall merging point P2of the second channel55along a cross section orthogonal to the second axis O2(hereinafter referred to as an overall merging point cross-sectional shape) is rectangular. A length L21of the overall merging point cross-sectional shape in the thickness direction Z is longer than the length L17of the first channel intermediate portion cross-sectional shape in the thickness direction Z. A length L22of the overall merging point cross-sectional shape in the orthogonal direction Y is equivalent to the length L18of the first channel intermediate portion cross-sectional shape in the orthogonal direction Y.

As shown inFIG.1, for example, an opening51communicates with the portions of the bent portions44and49that communicate with each other. For example, the opening51is formed in the channel substrate13and the protective substrate17.

The observation section60is formed at an end of the second channel55on the downstream side X2. In other words, the observation section60is provided on the side farther away from the first channel25than the overall merging point P2in the second channel55.

As shown inFIG.7, for example, the observation section60is formed through all the channel substrates11,12, and13. The observation section60transmits laser light (electromagnetic waves) L1from outside the observation chip1to the second channel55. That is, the portions of the protective substrates16and17corresponding to the observation section60are transparent. In this example, the laser light L1passes through the observation section60in the thickness direction Z. It should be noted that the protective substrate16may not be transparent. In this case, the observation section reflects the laser light L1for observation. The electromagnetic waves are not limited to the laser light L1, and may also be ultraviolet light or the like.

The length L21of the second channel55in the thickness direction Z is constant from the overall merging point P2to the observation section60.

The length of the second channel55in the orthogonal direction Y from the overall merging point P2to the observation section60gradually decreases as it goes from the overall merging point P2toward the downstream side X2, and then gradually decreases as it goes toward the downstream side X2. That is, the length of the second channel55in the orthogonal direction Y at the intermediate portion between the overall merging point P2and the observation section60is shorter than the length of the second channel55in the orthogonal direction Y at the overall merging point P2.

It should be noted that the length of the second channel55in the orthogonal direction Y from the overall merging point P2to the observation section60may be constant.

As shown inFIG.1, the third channel65extends from the observation section60toward the downstream side X2along the second axis O2. In this example, the length of the third channel65in the orthogonal direction Y is longer than the length L22of the second channel55(observation section60) in the orthogonal direction Y.

The separation section70is formed in a columnar shape. The separation section70is provided at an end of the third channel65on the downstream side X2.

For example, the second channel55, the observation section60, the third channel65, and the separation section70are respectively formed in the channel substrate12.

An opening71communicates with the separation section70. For example, the opening71is formed in the channel substrate13and the protective substrate17.

Next, the operation of the observation chip1configured as described above will be illustrated.

When using the observation chip1, a sample fluid supply tube (not shown) is connected to the opening21, and protective fluid tubes (not shown) are connected to the openings36and51, respectively. A sample fluid supply pump (not shown) is connected to the sample fluid supply tube, and a protective fluid supply pump (not shown) is connected to the protective fluid tubes.

First, the sample fluid supply pump is driven to supply a sample fluid containing cells, such as serum, to the first channel25through the sample fluid supply tube and the opening21.

Then, the protective fluid supply pump is driven to supply a protective fluid to the first branch channels31A and31B and the second branch channels41and46of the surrounding channel30through the protective fluid tubes and the openings36and51. The protective fluid is, for example, water, saline, or the like.

It should be noted that the sample fluid supply pump may be driven after the protective fluid supply pump is driven, or both supply pumps may be driven simultaneously.

InFIG.2toFIG.6, a white arrow B1schematically represents the flow of the sample fluid, and a white circle C1schematically represents the sample fluid. A gray (hatched) arrow B2schematically represents the flow of the protective fluid through the first branch channels31A and31B, and a gray circle C2schematically represents the protective fluid passing through the first branch channels31A and31B. A black arrow B3schematically represents the flow of the protective fluid through the second branch channels41and46, and a black circle C3schematically represents the protective fluid passing through the second branch channels41and46.

When both supply pumps continue to be driven, the sample fluid flows through the first channel25toward the downstream side X2, and the protective fluid flows through the first branch channels31A and31B and the second branch channels41and46toward the downstream side X2. As shown inFIG.4, at the partial merging point P1of the first channel25, the sample fluid C1is sandwiched from the orthogonal direction Y by the protective fluid C2passing through the first branch channels31A and31B. Further, as shown inFIG.5, as the length L18of the first channel intermediate portion cross-sectional shape in the orthogonal direction Y is shorter than the length L15of the partial merging point cross-sectional shape, the sample fluid C1is confined between the protective fluid C2at the intermediate portion between the partial merging point P1and the overall merging point P2.

Then, as shown inFIG.6, at the overall merging point P2, the sample fluid C1and the protective fluid C2are sandwiched from the thickness direction Z by the protective fluid C3passing through the second branch channels41and46.

In the observation chip1, as described above, after the sample fluid C1is sandwiched by the protective fluid C2in the orthogonal direction Y, the sample fluid C1and the protective fluid C2are sandwiched by the protective fluid C3in the thickness direction Z.

In this way, the protective fluids C2and C3surround the sample fluid C1all around. The sample fluid C1surrounded by the protective fluids C2and C3flows through the second channel55, and is then observed using the laser light L1in the observation section60.

The observed sample fluid C1and protective fluids C2and C3flow into the separation section70through the third channel65. The sample fluid C1and the protective fluids C2and C3flowing into the separation section70are processed as appropriate.

Here, the flow of the sample fluid C1in the second channel55of the observation chip1will be illustrated.

As shown inFIG.8, in the observation chip1, the length of the second channel55in the thickness direction Z is constant from the overall merging point P2to the observation section60. Therefore, the flow of the sample fluid C1sandwiched by the protective fluid C3is less likely to be disturbed.

In contrast,FIG.9shows a cross-sectional view of an observation chip2in the related art, such as Non-Patent Document 1. The second channel75of the observation chip2is formed with step portions76and77that narrow the channel in the thickness direction Z. Therefore, the flow of the sample fluid C1and the protective fluid C3is disturbed at the step portions76and77, making it difficult to surround the sample fluid C1with the protective fluids C2and C3. Thus, it becomes difficult to observe the sample fluid C1using the laser light L1or the like in the observation section80.

Next, a simulation result of the flow of the sample fluid C1in the second channel55of the observation chip1will be illustrated.

As shown inFIG.10, it can be seen that the flow of the sample fluid C1surrounded by the protective fluids C2and C3is stable without disturbance along the second axis O2in the second channel55.

As illustrated above, in the observation chip1of this embodiment, the sample fluid C1flows from the first channel25toward the overall merging point P2, and the protective fluids C2and C3flow from the surrounding channel30toward the overall merging point P2. At this time, the sample fluid C1is surrounded by the protective fluids C2and C3all around in the cross section S2. Since the length of the second channel55in the thickness direction Z is constant from the overall merging point P2to the observation section60, the flow of the sample fluid C1and the protective fluids C2and C3is not disturbed. Thus, the observation chip1is configured to stabilize the sample fluid C1for observation in the observation section60.

The surrounding channel30includes the first branch channels31A and31B and the second branch channels41and46. Therefore, the sample fluid C1may be sandwiched at once from the orthogonal direction Y by the protective fluid C2flowing through the first branch channels31A and31B, and the sample fluid C1and the protective fluid C2may be sandwiched at once from the thickness direction Z by the protective fluid C3flowing through the second branch channels41and46. The first branch channels31A and31B are plane symmetrical with respect to the reference plane S1. Therefore, the sample fluid C1may be stably sandwiched by the protective fluid C2flowing through the first branch channels31A and31B.

The second branch channels41and46may be plane symmetrical with respect to the reference plane S1. In this case, the sample fluid C1may be stably sandwiched by the protective fluid C3flowing through the second branch channels41and46.

The length L18of the first channel25and the first branch channels31A and31B in the orthogonal direction Y at the intermediate portion between the partial merging point P1and the overall merging point P2is shorter than the length L15of the first channel25and the first branch channels31A and31B in the orthogonal direction Y at the partial merging point P1. Thereby, the sample fluid C1flowing through the first channel25may be more reliably confined between the protective fluid C2flowing through the first branch channels31A and31B.

The length of the second channel55in the orthogonal direction Y at the intermediate portion between the overall merging point P2and the observation section60is shorter than the length L22of the second channel55in the orthogonal direction Y at the overall merging point P2. Therefore, the performance of confining the sample fluid C1with the protective fluids C2and C3as a whole may be improved.

It should be noted that an observation chip3according to a modified example may be configured as shown inFIG.11andFIG.12. In the observation chip3, as shown inFIG.11, the sample fluid C1is sandwiched by the protective fluid C3from the thickness direction Z at a partial merging point P6. Thereafter, as shown inFIG.12, the sample fluid C1and the protective fluid C3are sandwiched by the protective fluid C2from the orthogonal direction Y at an overall merging point P7.

The observation chip3of the modified example is configured to achieve the same effects as the observation chip1of this embodiment.

An embodiment of the present disclosure has been described in detail above with reference to the drawings, but the specific configuration is not limited to this embodiment, and the present disclosure includes modifications, combinations, deletions, etc. of the configuration without departing from the gist of the present disclosure.

For example, in the above embodiment, the first branch channels31A and31B are not necessarily plane symmetrical with respect to the reference plane S1.

It should be noted that a sorting device for sorting cells, for example, also needs to have a structure in which particles stably flow at a fixed position in the channel in a cross section orthogonal to the channel, as used in this embodiment.