阅读说明：本技术 试样支撑体 (Sample support ) 是由 小谷政弘 大村孝幸 于 2020-01-23 设计创作，主要内容包括：本发明的试样支撑体是用于试样的电离的试样支撑体,具备：基板,其具有第一表面、及与第一表面为相反侧的第二表面、以及在第一表面及第二表面的各个开口的多个贯通孔；框架,其安装于基板,框架的热传导率为1.0W/m·K以下。(The sample support of the present invention is a sample support for ionizing a sample, and includes: a substrate having a first surface, a second surface opposite to the first surface, and a plurality of through holes opened in the first surface and the second surface, respectively; and a frame attached to the substrate, wherein the thermal conductivity of the frame is 1.0W/mK or less.)

1. A sample support body, wherein,

is a sample support for ionization of a sample,

the disclosed device is provided with:

a substrate having a first surface, a second surface opposite to the first surface, and a plurality of through holes opened in the first surface and the second surface, respectively; and

a frame mounted to the substrate,

the thermal conductivity of the frame is 1.0W/mK or less.

2. The sample support according to claim 1,

the width of each of the through holes is 1 to 700nm,

the thickness of the substrate is 1-50 μm.

3. The sample support according to claim 1 or 2, wherein,

the substrate is formed by anodizing a valve metal or silicon.

4. The sample support according to any one of claims 1 to 3, wherein,

the substrate and the frame are each made of an electrically insulating material.

5. The sample support according to claim 4,

the frame is made of ceramic or glass.

6. The sample support according to claim 4,

the material of the frame is resin.

7. The sample support according to claim 6,

the resin is PET, PEN or PI.

8. The sample support according to any one of claims 1 to 7, wherein,

the thickness of the frame is 10-500 μm.

9. The sample support according to any one of claims 1 to 8, wherein,

the frame is transmissive to visible light.

10. The sample support according to any one of claims 1 to 9, wherein,

the frame has flexibility.

11. The sample support according to any one of claims 1 to 10, wherein,

the substrate is a plurality of substrates,

the frame is a plurality of frames corresponding to the plurality of substrates,

the plurality of frames are connected to each other in a state of being arranged in at least 1 column.

Technical Field

The present disclosure relates to a sample support.

Background

Examples of Ionization methods for ionizing a sample such as a biological sample for mass analysis include Matrix-Assisted Laser Desorption Ionization (MALDI), Surface-Assisted Laser Desorption/Ionization (SALDI), and Desorption Electrospray Ionization (DESI). The matrix-assisted laser desorption ionization method is a method in which a low-molecular-weight organic compound called a matrix that absorbs laser light is added to a sample, and the sample is ionized by irradiating the sample with laser light. The surface-assisted laser desorption ionization method is a method in which a sample is dropped onto an ionization substrate having a fine uneven structure on the surface thereof, and the sample is irradiated with a laser beam to ionize the sample. The desorption electrospray ionization method is a method of irradiating a sample with charged fine droplets (charged-droplets) to desorb and ionize the sample.

As a sample support capable of ionizing a component of a sample while maintaining positional information of the component of the sample (information on a two-dimensional distribution of molecules constituting the sample), there is known a sample support including a substrate having a first surface, a second surface opposite to the first surface, and a plurality of through holes opened in the first surface and the second surface (see, for example, patent document 1). In such a sample support, when the second surface of the substrate is brought into contact with the sample, the components of the sample move from the second surface side to the first surface side through the plurality of through holes in the substrate, and stay on the first surface side.

Documents of the prior art

Patent document

Patent document 1: japanese patent No. 6093492

Disclosure of Invention

Problems to be solved by the invention

In the ionization method as described above, a frozen sample is often used as a target. In this case, in the sample support as described above, it is very important how components of the sample can be uniformly moved through the plurality of through-holes.

An object of the present disclosure is to provide a sample support body that can uniformly move components of a sample through a plurality of through holes, particularly when a frozen sample is used.

Means for solving the problems

One aspect of the present disclosure provides a sample support for ionization of a sample, including: a substrate having a first surface, a second surface opposite to the first surface, and a plurality of through holes opened in the first surface and the second surface, respectively; and a frame attached to the substrate, wherein the thermal conductivity of the frame is 1.0W/mK or less.

In this sample support, the second surface of the substrate is brought into contact with the frozen sample, and when the sample is thawed in this state, components of the sample move from the second surface side to the first surface side in the substrate through the plurality of through holes and stay on the first surface side. At this time, since the thermal conductivity of the frame is 1.0W/m · K or less, even if the frame is handled by hand, for example, heat conduction to the sample through the frame can be suppressed, and as a result, thawing of the sample is performed uniformly. When the thawing of the sample is performed uniformly, the sample is brought into uniform contact with the second surface of the substrate, and as a result, the components of the sample are reliably moved from the second surface side to the first surface side via the plurality of through holes. Therefore, according to the sample support, particularly when a frozen sample is used, the components of the sample can be uniformly moved through the plurality of through holes.

In the sample support according to one aspect of the present disclosure, the width of each of the plurality of through holes may be 1 to 700nm, and the thickness of the substrate may be 1 to 50 μm. Thus, when the frozen sample is brought into contact with the second surface of the substrate to thaw the sample, the components of the sample can smoothly move from the second surface side to the first surface side through the plurality of through holes in the substrate and stay on the first surface side in an appropriate state.

In the sample support according to one aspect of the present disclosure, the substrate may be formed by anodizing the valve metal or silicon. This makes it possible to easily and reliably obtain a substrate having a plurality of through holes.

In the sample support according to one aspect of the present disclosure, the material of each of the substrate and the frame may be an electrically insulating material. Thus, for example, in the desorption electrospray ionization method, even if the fine droplet irradiation portion to which a high voltage is applied is close to the first surface, the generation of discharge between the fine droplet irradiation portion and the sample support can be suppressed. Therefore, in the desorption electrospray ionization method, particularly in the case of using a frozen sample, the components of the sample can be reliably ionized by irradiation of the charged minute droplets.

In the sample support of one aspect of the present disclosure, the material of the frame may also be ceramic or glass. Thus, an electrically insulating frame having a thermal conductivity of 1.0W/mK or less can be easily obtained. In particular, if the material of the frame is ceramic or glass, the shrinkage of the sample can be suppressed when the frozen sample is thawed.

In the sample support of one aspect of the present disclosure, the material of the frame may also be a resin. Thus, an electrically insulating frame having a thermal conductivity of 1.0W/mK or less can be easily obtained. In the sample support of one aspect of the present disclosure, the resin may also be PET, PEN, or PI. This makes it possible to more easily obtain an electrically insulating frame having a thermal conductivity of 1.0W/mK or less.

In the sample support according to one aspect of the present disclosure, the frame may have a thickness of 10 to 500 μm. Thus, for example, in the desorption electrospray ionization method, even if the fine droplet irradiation portion is close to the first surface, physical interference between the fine droplet irradiation portion and the frame is not likely to occur. Therefore, in the desorption electrospray ionization method, the charged fine droplets are irradiated onto the first surface by bringing the fine droplet irradiation portion close to the first surface, so that the components of the sample moving to the first surface side through the plurality of through holes can be reliably ionized.

In the sample support of one aspect of the present disclosure, the frame may also have a transmissivity with respect to visible light. This improves the visibility of the sample through the frame, and therefore, the second surface of the substrate can be reliably brought into contact with the sample.

In the sample support according to one aspect of the present disclosure, the frame may have flexibility. This can improve the ease of handling the sample support.

In the sample support according to one aspect of the present disclosure, the substrate may be a plurality of substrates, the frame may be a plurality of frames corresponding to the plurality of substrates, and the plurality of frames may be connected to each other in a state of being arranged in at least 1 row. This enables the corresponding substrate and frame to be separated by a necessary amount and used.

Effects of the invention

According to the present disclosure, a sample support capable of uniformly moving a component of a sample through a plurality of through holes can be provided, particularly when a frozen sample is used.

Drawings

Fig. 1 is a plan view of a sample support according to an embodiment.

FIG. 2 is a sectional view of the sample support taken along line II-II shown in FIG. 1.

Fig. 3 is an enlarged image of the substrate of the sample support shown in fig. 1.

Fig. 4 is a diagram showing a process of a mass spectrometry method using the sample support shown in fig. 1.

Fig. 5 is a diagram showing a process of a mass spectrometry method using the sample support shown in fig. 1.

Fig. 6 is a block diagram of a mass spectrometer for performing a mass spectrometry method using the sample support shown in fig. 1.

Fig. 7 is a perspective view of a sample support according to a modification.

Fig. 8 is a diagram showing a process of a mass spectrometry method using a sample support according to a modification.

Detailed Description

Hereinafter, embodiments of the present disclosure will be described in detail with reference to the accompanying drawings. In the drawings, the same or corresponding portions are denoted by the same reference numerals, and redundant description thereof will be omitted.

[ sample support ]

As shown in fig. 1 and 2, the sample support 1 includes a substrate 2, a frame 3, and an adhesive layer 4. The substrate 2 has a first surface 2a, a second surface 2b, and a plurality of through holes 2 c. The second surface 2b is a surface opposite to the first surface 2 a. The through holes 2c are opened in the first surface 2a and the second surface 2b, respectively. In the present embodiment, a plurality of through holes 2c are formed in the entire substrate 2 in the same manner (with uniform distribution), and each through hole 2c extends in the thickness direction of the substrate 2 (the direction in which the first surface 2a and the second surface 2b face each other).

The substrate 2 is an electrically insulating member. In the present embodiment, the thickness of the substrate 2 is 1 to 50 μm, and the width of each through hole 2c is about 1 to 700 nm. The shape of the substrate 2 when viewed from the thickness direction of the substrate 2 is, for example, a substantially circular shape having a diameter of about several mm to several cm. The through-holes 2c are, for example, substantially circular when viewed in the thickness direction of the substrate 2 (see fig. 3). The width of the through-hole 2c refers to the diameter of the through-hole 2c when the shape of the through-hole 2c is circular when viewed from the thickness direction of the substrate 2, and refers to the diameter (effective diameter) of a virtual maximum cylinder that is received in the through-hole 2c when the shape is other than circular.

The frame 3 has a third surface 3a and a fourth surface 3b, and an opening 3 c. The fourth surface 3b is a surface on the opposite side to the third surface 3a, i.e., a surface on the substrate 2 side. The opening 3c is opened in each of the third surface 3a and the fourth surface 3 b. The frame 3 is an electrically insulating member, and the thermal conductivity of the frame 3 is 1.0W/m.K or less. In the present embodiment, the material of the frame 3 is PET (polyethylene terephthalate), PEN (polyethylene naphthalate), or PI (polyimide), and the thickness of the frame 3 is 10 to 500 μm (more preferably 100 μm or less). In the present embodiment, the frame 3 has transmissivity with respect to visible light, and the frame 3 has flexibility. The frame 3 has a rectangular shape, for example, with one side of the rectangular shape being several cm, when viewed from the thickness direction of the substrate 2. The shape of the opening 3c when viewed from the thickness direction of the substrate 2 is, for example, a circle having a diameter of about several mm to several cm. The lower limit of the thermal conductivity of the frame 3 is, for example, 0.1W/m.K.

The frame 3 is attached to the substrate 2. In the present embodiment, the region along the outer edge of the substrate 2 in the first surface 2a of the substrate 2 and the region along the outer edge of the opening 3c in the fourth surface 3b of the frame 3 are fixed to each other by the adhesive layer 4. The material of the adhesive layer 4 is, for example, an adhesive material (low melting point glass, vacuum adhesive, or the like) which releases little gas. In the sample support 1, a portion of the substrate 2 corresponding to the opening 3c of the frame 3 functions as an effective region R for moving a component of the sample from the second surface 2b side to the first surface 2a side through the plurality of through holes 2 c.

Fig. 3 is an enlarged image of the substrate 2 when viewed from the thickness direction of the substrate 2. In fig. 3, black portions are through holes 2c, and white portions are partition walls between the through holes 2 c. As shown in fig. 3, a plurality of through holes 2c having a substantially constant width are formed in the substrate 2 in a uniform manner. The aperture ratio of the through holes 2c in the effective region R (the ratio of all the through holes 2c to the effective region R when viewed in the thickness direction of the substrate 2) is practically 10 to 80%, and particularly preferably 60 to 80%. The sizes of the plurality of through holes 2c may be different from each other, or the plurality of through holes 2c may be partially connected to each other.

The substrate 2 shown in fig. 3 is an alumina porous film formed by anodizing Al (aluminum). Specifically, the Al substrate is anodized, and the oxidized surface portion is peeled off from the Al substrate, whereby the substrate 2 can be obtained. The substrate 2 may be formed by anodizing a valve metal other than Al, such as Ta (tantalum), Nb (niobium), Ti (titanium), Hf (hafnium), Zr (zirconium), Zn (zinc), W (tungsten), Bi (bismuth), and Sb (antimony), or may be formed by anodizing Si (silicon).

[ ionization method and Mass analysis method ]

An ionization method and a mass analysis method using the sample support 1 will be described. The ionization method here is desorption electrospray ionization. Since the desorption electrospray ionization method is performed in an atmospheric pressure atmosphere, it is advantageous in that the sample can be directly analyzed, and the sample can be easily exchanged and observed and analyzed. In fig. 4 and 5, the through-hole 2c and the adhesive layer 4 are not shown in the sample support 1. For convenience of illustration, the sample support 1 shown in fig. 1 and 2 and the sample support 1 shown in fig. 4 and 5 are different in the ratio of the dimensions and the like.

First, the above-described sample support 1 is prepared as a sample support for sample ionization (first step). The sample support 1 may be prepared by manufacturing by a manufacturer of the ionization method and the mass spectrometry method, or may be prepared by transferring from a manufacturer, a seller, or the like of the sample support 1.

Next, as shown in fig. 4 (a), the sample S is placed on the placement surface 6a of the slide glass (placement portion) 6 (second step). The sample S is a thin film-like biological sample (aqueous sample) such as a tissue slice, and is in a frozen state. Next, as shown in fig. 4 (b), the sample support 1 is placed on the placement surface 6a so that the second surface 2b of the substrate 2 is in contact with the sample S (second step). At this time, the sample support 1 is disposed so that the sample S is located within the effective region R when viewed from the thickness direction of the substrate 2. Next, as shown in fig. 5 (a), the frame 3 is fixed to the slide glass 6 using an electrically insulating tape 7. When the sample S is thawed in this state, as shown in fig. 5 (b), the component S1 of the sample S moves from the second surface 2b side to the first surface 2a side through the plurality of through holes 2c (see fig. 2) by, for example, capillary action in the substrate 2, and the component S1 of the sample S stays on the first surface 2a side by, for example, surface tension.

Next, if the sample S is dried, as shown in fig. 6, the slide glass 6, the sample S, and the sample support 1 are placed on the stage 21 in the ionization chamber 20 of the mass spectrometer 10. The ionization chamber 20 is filled with an atmospheric pressure atmosphere. Next, the charged fine droplets I are irradiated onto the region corresponding to the effective region R on the first surface 2a of the substrate 2, whereby the component S1 of the sample S moving to the first surface 2a side is ionized, and sample ions S2 as the ionized component are attracted (third step). In the present embodiment, for example, by moving the stage 21 in the X-axis direction and the Y-axis direction, the irradiation region I1 of the charged fine droplet I is moved relative to the region corresponding to the effective region R on the first surface 2a of the substrate 2 (that is, the charged fine droplet I is scanned over the region). The first step, the second step, and the third step described above correspond to the desorption electrospray ionization method using the sample support 1.

In the ionization chamber 20, the charged fine droplets I are ejected from the nozzle 22, and the sample ions S2 are attracted from the suction port of the ion transport tube 23. The nozzle 22 has a double-layered cartridge structure. The solvent is guided to the inner cylinder of the nozzle 22 in a state where a high voltage is applied. This gives a biased charge to the solvent reaching the tip of the nozzle 22. The atomizing gas is directed toward the outer barrel of the nozzle 22. Thereby, the solvent is atomized into fine droplets, and solvent ions generated in the process of vaporizing the solvent are emitted as charged fine droplets I.

The sample ions S2 sucked through the suction port of the ion transport tube 23 are transported through the ion transport tube 23 into the mass spectrometer 30. The inside of the mass analysis chamber 30 is in a high vacuum atmosphere (degree of vacuum 10)^-4An atmosphere of below Torr). In the mass analysis chamber 30, the sample ions S2 are converged by the ion optical system 31 and introduced into the quadrupole mass filter 32 to which a high-frequency voltage is applied. When the sample ions S2 are introduced into the quadrupole mass filter 32 to which the high-frequency voltage is applied, ions having a mass number determined in accordance with the frequency of the high-frequency voltage are selectively passed, and the passed ions are detected by the detector 33 (fourth step). The mass number of ions reaching the detector 33 is sequentially changed by sweeping the frequency of the high-frequency voltage applied to the quadrupole mass filter 32, and a mass spectrum in a predetermined mass range is obtained. In the present embodiment, the detector 33 detects ions so as to correspond to the position of the irradiation region I1 of the charged fine droplet I, and images the two-dimensional distribution of the molecules constituting the sample S. The above first step, second step, third step and fourth step correspond to a mass spectrometry method using the sample support 1.

[ Effect and Effect ]

When the second surface 2b of the substrate 2 is brought into contact with the frozen sample S in the sample support 1 and the sample S is thawed in this state, the component S1 of the sample S moves from the second surface 2b side to the first surface 2a side through the plurality of through holes 2c in the substrate 2 and stays on the first surface 2a side. At this time, since the thermal conductivity of the frame 3 is 1.0W/m · K or less, even if the frame 3 is handled by hands, for example, heat conduction to the sample S through the frame 3 can be suppressed, and as a result, thawing of the sample S is performed uniformly. When the thawing of the sample S is uniformly performed, the sample S and the second surface 2b of the substrate 2 are uniformly brought into contact with each other, and as a result, the component S1 of the sample S reliably moves from the second surface 2b side to the first surface 2a side through the plurality of through holes 2 c. Therefore, according to the sample support 1, particularly when the frozen sample S is used, the component S1 of the sample S can be uniformly moved through the plurality of through-holes 2 c.

In the sample support 1, the width of each through-hole 2c is 1 to 700nm, and the thickness of the substrate 2 is 1 to 50 μm. Thus, when the second surface 2b of the substrate 2 is brought into contact with the frozen sample S and the sample S is thawed in this state, the component S1 of the sample S can smoothly move from the second surface 2b side to the first surface 2a side through the plurality of through holes 2c in the substrate 2 and can stay on the first surface 2a side in an appropriate state.

In the sample support 1, the substrate 2 is formed by anodizing a valve metal or silicon. This makes it possible to easily and reliably obtain the substrate 2 having the plurality of through holes 2 c.

In the sample support 1, the material of each of the substrate 2 and the frame 3 is an electrically insulating material. Thus, for example, in the desorption electrospray ionization method, even if the nozzle 22, which is a fine droplet irradiation portion to which a high voltage is applied, is close to the first surface 2a, the generation of discharge between the nozzle 22 and the sample support 1 can be suppressed. When the distance between the nozzle 22 and the sample support 1 is shortened, the spread of electrospray (spray of charged minute droplets) is suppressed in image formation, and therefore, the spatial resolution can be improved. Therefore, as described above, it is extremely effective to make the nozzle 22 close to the first surface 2a in order to reliably ionize the component S1 of the sample S. Therefore, in the desorption electrospray ionization method, particularly when a frozen sample S is used, the component S1 of the sample S can be reliably ionized by irradiation of the charged fine droplets I.

In the sample support 1, the material of the frame 3 is PET, PEN, or PI. This makes it possible to easily obtain an electrically insulating frame 3 having a thermal conductivity of 1.0W/mK or less.

In the sample support 1, the frame 3 has a thickness of 10 to 500 μm (more preferably 100 μm or less). Thus, for example, in the desorption electrospray ionization method, even if the nozzle 22 is close to the first surface 2a, physical interference between the nozzle 22 and the frame 3 is not easily generated. Therefore, in the desorption electrospray ionization method, the component S1 of the sample S moving to the first surface 2a side via the plurality of through holes 2c can be reliably ionized by bringing the nozzle 22 close to the first surface 2a and irradiating the first surface 2a with the charged fine droplets I.

In the sample support 1, the frame 3 has transparency to visible light. This improves the visibility of the sample S through the frame 3, and therefore, the second surface 2b of the substrate 2 can be reliably brought into contact with the sample S.

In the sample support 1, the frame 3 has flexibility. This can improve the ease of handling the sample support 1.

[ modified examples ]

The present disclosure is not limited to the above-described embodiments. For example, as shown in fig. 7, the sample support 1 may include a plurality of substrates 2 and a plurality of frames 3 corresponding to the plurality of substrates 2, and the plurality of frames 3 may be connected to each other in a state of being arranged in at least 1 row. This enables the corresponding substrate 2 and frame 3 to be separated by a necessary amount and used. In this case, if the frames 3 have flexibility, the sample support 1 can be processed in a state in which the plurality of frames 3 connected to each other in a state of being arranged in at least 1 row are wound in a roll shape.

The material of the frame 3 may be a resin other than PET, PEN, or PI. In this case, the frame 3 having thermal conductivity of 1.0W/mK or less can be easily obtained. The material of the frame 3 may be ceramic or glass. In this case, the frame 3 having thermal conductivity of 1.0W/mK or less can be easily obtained. In particular, if the material of the frame 3 is ceramic or glass, the sample S can be suppressed from shrinking when the frozen sample S is thawed. Further, if the frame 3 having a thermal conductivity of 0.1W/m · K can be realized, the material of the frame 3 is not particularly limited. The frame 3 may be colored by, for example, a pigment. This enables the sample support 1 to be classified according to the application.

In the above-described embodiment, one effective region R is provided on the substrate 2, but a plurality of effective regions R may be provided on the substrate 2. In the above-described embodiment, the plurality of through holes 2c are formed in the entire substrate 2, but the plurality of through holes 2c may be formed in a portion of the substrate 2 corresponding to at least the effective region R. In the above-described embodiment, the sample S is arranged so that one sample S corresponds to one effective region R, but a plurality of samples S may be arranged so that one effective region R corresponds to the sample S.

Alternatively, an opening different from the opening 3c may be provided in the frame 3, and the sample support 1 may be fixed to the slide glass 6 with the tape 7 through the opening. The sample support 1 may be fixed to the slide glass 6 by means other than the tape 7 (for example, means using an adhesive, a fixing member, or the like). For example, as shown in fig. 8, the sample support 1 may be fixed to the slide glass 6 using a gel 8. In this case, the gel 8 is preferably a material (for example, glycerin or the like) that does not solidify in a low-temperature environment in which the frozen sample S is processed. As a step, the gel 8 is applied to the region (for example, four corners of the frame 3) of the surface of the frame 3 on the substrate 2 side where the substrate 2 is not fixed. At this time, the gel 8 is applied to the effective region R of the substrate 2 so that the gel 8 does not overflow the region. Next, the effective region R of the substrate 2 is brought into contact with the sample S, and the sample support 1 is placed on the placement surface 6a of the slide glass 6. In addition, when the material of the frame 3 is resin, the sample support 1 can be fixed to the slide glass 6 by static electricity.

The sample S is not limited to a water-containing sample, and may be a dry sample. When the sample S is a dry sample, a solution (e.g., acetonitrile mixture solution) for reducing the viscosity of the sample S is added to the sample S. Thus, for example, the component S1 of the sample S can be moved toward the first surface 2a of the substrate 2 through the plurality of through holes 2c by capillary action.

The sample support 1 may be used for other ionization methods than the desorption electrospray ionization method. In other ionization methods, the frame 3 may sometimes have conductivity. In other ionization methods, the substrate 2 itself may have conductivity, or a conductive film may be formed on the substrate 2. The material of the conductive film is preferably a metal having low affinity with a sample (e.g., protein), and is preferably Au (gold), Pt (platinum), Cr (chromium), Ni (nickel), Ti (titanium), or the like.

The structures of the above-described embodiments are not limited to the materials and shapes described above, and various materials and shapes can be applied. The configurations of the above-described one embodiment or modification can be arbitrarily applied to the configurations of the other embodiments or modifications.

Description of the symbols

1 … … sample support; 2 … … a substrate; 2a … … first surface; 2b … … second surface; 2c … … through holes; 3 … … frame.