Patent Publication Number: US-2022238317-A1

Title: Sample support, sample ionization method, and mass spectrometry method

Description:
TECHNICAL FIELD 
     The present disclosure relates to a sample support, a sample ionization method, and a mass spectrometry method. 
     BACKGROUND ART 
     In the related art, a sample support for ionizing a sample is known in mass spectrometry of a sample such as a biological sample (for example, refer to Patent Literature 1). Such a sample support includes a substrate formed with a plurality of through holes opening to a first surface and a second surface on a side opposite to the first surface. In a case where the sample support is disposed on the sample such that the second surface faces the sample, it is possible to lift up the sample from the second surface side of the substrate toward the first surface side through the through hole by using a capillary action. Then, in a case where the first surface side, for example, is irradiated with an energy ray such as laser beam, the sample moved to the first surface side is ionized. 
     CITATION LIST 
     Patent Literature 
     Patent Literature 1: Japanese Patent No. 6093492 
     SUMMARY OF INVENTION 
     Technical Problem 
     In the mass spectrometry as described above, when the first surface side of the substrate is irradiated with the energy ray, it is required that a mass spectrometry device recognizes an irradiation range of the energy ray. However, a visual field of a camera or the like that is attached to the mass spectrometry device is narrow in accordance with the mass spectrometry device, and it is not possible to observe the entire sample support disposed in the mass spectrometry device, and thus, it may not be possible to easily recognize the irradiation range. 
     Therefore, an object of one aspect of the present disclosure is to provide a sample support, a sample ionization method, and a mass spectrometry method in which an irradiation range of an energy ray can be easily recognized. 
     Solution to Problem 
     A sample support according to one aspect of the present disclosure is a sample support for sample ionization, including: a substrate formed with a plurality of through holes opening to a first surface and a second surface on a side opposite to the first surface; a conductive layer provided not to block the through hole in the first surface; and a frame body provided in a peripheral portion of the substrate to surround an ionization region in which a sample is ionized when viewed in a thickness direction of the substrate, in which a marker for recognizing a position in the ionization region is provided in the frame body. 
     In the sample support, the plurality of through holes opening to the first surface and the second surface on a side opposite to the first surface are formed on the substrate. For this reason, for example, in a case where the sample support is disposed on a sample such as a biological sample such that the second surface of the substrate faces the sample, it is possible to move the sample (a component of the sample) toward the first surface side from the second surface side through the through hole by using a capillary action. Further, for example, in a case where the first surface is irradiated with an energy ray such as laser beam, energy is transmitted to the component of the sample moved to the first surface side via the conductive layer, and thus, it is possible to ionize the component of the sample. In addition, the sample support includes the frame body provided in the peripheral portion of the substrate. For this reason, it is possible to improve the handleability of the sample support by the frame body. In addition, the frame body surrounds the ionization region in which the sample is ionized when viewed in the thickness direction of the substrate, and the marker for recognizing the position in the ionization region is provided in the frame body. Accordingly, the following effects are obtained. That is, for example, a visual field of a camera or the like that is attached to an ionization device irradiating the sample support with an energy ray is narrow, and it may be difficult to specify an irradiation range (a range to be irradiated with the energy ray) by the observation of the ionization region. Even in such a case, it is possible for the ionization device to recognize the irradiation range of the energy ray, by causing the camera or the like to perform scanning and by reading the marker provided in the frame body. Accordingly, according to such a sample support, it is possible to easily recognize the irradiation range of the energy ray. 
     A width of the through hole may be 1 nm to 700 nm, and a thickness of the substrate may be 1 μm to 50 μm. In this case, it is possible to suitably attain the movement of the component of the sample by the capillary action described above. 
     A plurality of first markers disposed along a first direction may be provided in a portion of the frame body extending along the first direction, and a plurality of second markers disposed along a second direction orthogonal to the first direction may be provided in a portion of the frame body extending along the second direction. In this case, it is possible to recognize the position in the first direction by the first marker and to recognize the position in the second direction by the second marker. Accordingly, it is possible to easily grasp two-dimensional coordinates of the irradiation range of the energy ray (for example, a start point position, an end point position, and the like). 
     The marker may be at least one selected from a numeric character, a signal, and a letter. In this case, it is possible to attain the marker suitable for visual contact and/or for reading a device. 
     The marker may include a marker for visual contact having a width of greater than or equal to a predetermined value and a marker for a device having a width of less than the predetermined value. In this case, for example, it is possible for a measurer to determine in advance the irradiation range by visually reading the marker for visual contact. Further, for example, the marker for a device corresponding to the irradiation range determined by the measurer is read by the camera that is attached to the ionization device, and thus, it is possible for the ionization device to recognize the irradiation range of the energy ray. 
     A sample support according to another aspect of the present disclosure is a sample support for sample ionization, including: a substrate having conductivity, and formed with a plurality of through holes opening to a first surface and a second surface on a side opposite to the first surface; and a frame body provided in a peripheral portion of the substrate to surround an ionization region in which a sample is ionized when viewed in a thickness direction of the substrate, in which a marker for recognizing a position in the ionization region is provided in the frame body. 
     According to such a sample support, it is possible to omit the conductive layer and to obtain the same effects as those of the sample support including the conductive layer described above. 
     A sample ionization method according to one aspect of the present disclosure is a sample ionization method of an ionization device including an irradiation unit configured to apply an energy ray, a scanning unit configured to scan a marker provided in a frame body, and a control unit configured to control an operation of the irradiation unit, the method including: a first step of preparing a sample and the sample support including the conductive layer; a second step of disposing the sample support on the sample such that the second surface faces the sample; a third step of causing the control unit to recognize an irradiation range of the energy ray in the ionization region by causing the scanning unit to scan the marker provided in the frame body; and a fourth step of ionizing a component of the sample moved to the first surface side through the through hole in the irradiation range by causing the control unit to operate the irradiation unit such that the first surface in the irradiation range is irradiated with the energy ray while a voltage is applied to the conductive layer. 
     In the sample ionization method described above, the plurality of through holes opening to the first surface and the second surface on a side opposite to the first surface are formed on the substrate. In a case where the sample support is disposed on the sample such that the second surface of the substrate faces the sample, the sample (the component of the sample) is moved toward the first surface side from the second surface side through the through hole by a capillary action. Further, in a case where the first surface is irradiated with the energy ray while a voltage is applied to the conductive layer, energy is transmitted to the component of the sample moved to the first surface side. Accordingly, the component of the sample is ionized. In addition, it is possible for the ionization device to easily recognize the irradiation range of the energy ray by scanning the marker provided in the frame body. 
     A sample ionization method according to another aspect of the present disclosure is a sample ionization method of an ionization device including an irradiation unit configured to apply an energy ray, a scanning unit configured to scan a marker provided in a frame body, and a control unit configured to control an operation of the irradiation unit, the method including: a first step of preparing a sample and the sample support including the substrate having conductivity; a second step of disposing the sample support on the sample such that the second surface faces the sample; a third step of causing the control unit to recognize an irradiation range of the energy ray in the ionization region by causing the scanning unit to scan the marker provided in the frame body; and a fourth step of ionizing a component of the sample moved to the first surface side through the through hole in the irradiation range by causing the control unit to operate the irradiation unit such that the first surface in the irradiation range is irradiated with the energy ray while a voltage is applied to the substrate. 
     According to such a sample ionization method, it is possible to omit the conductive layer from the sample support and to obtain the same effects as those in the case of using the sample support including the conductive layer as described above. 
     In the ionization method described above, the marker may include a marker for visual contact having a width of greater than or equal to a predetermined value and a marker for a device having a width of less than the predetermined value, and in the third step, a measurer may determine the irradiation range, on the basis of an existence range of the sample in the ionization region and the marker for visual contact, and the control unit may recognize the irradiation range, on the basis of a position of the scanning unit when the marker for a device corresponding to the irradiation range determined by the measurer is read by the scanning unit. In this case, it is possible to accurately attain both of the determination of the irradiation range by the visual contact of the measurer and the recognition of the irradiation range by a mechanical manipulation (marker scanning) of the ionization device, by the marker provided in the frame body. 
     A mass spectrometry method, includes: each of the steps of the sample ionization method according to one aspect of the present disclosure described above; a fifth step of detecting the ionized component and of acquiring a distribution image indicating a mass distribution of the sample in the irradiation range; a sixth step of acquiring an optical image including the sample and the sample support, in a state in which the sample support is disposed on the sample; and a seventh step of superimposing the optical image on the distribution image such that the irradiation range of the optical image overlaps with the distribution image, on the basis of the marker in the optical image. 
     According to the mass spectrometry method described above, it is possible to accurately superimpose the optical image of the sample on the distribution image, on the basis of the marker provided in the frame body of the sample support. As a result thereof, it is possible to visualize the mass distribution in each position of the sample. 
     Advantageous Effects of Invention 
     According to one aspect of the present disclosure, it is possible to provide a sample support, a sample ionization method, and a mass spectrometry method in which an irradiation range of an energy ray can be easily recognized. 
    
    
     
       BRIEF DESCRIPTION OF DRAWINGS 
         FIG. 1  is a plan view of a sample support according to one embodiment. 
         FIG. 2  is a sectional view of the sample support along line II-II illustrated in  FIG. 1 . 
         FIG. 3  is a diagram illustrating an enlarged image of an effective region in a substrate viewed in a thickness direction of the substrate illustrated in  FIG. 1 . 
         FIG. 4  is an enlarged view of a frame illustrated in  FIG. 1 . 
         FIG. 5  is a diagram illustrating a procedure of a mass spectrometry method according to one embodiment. 
         FIG. 6  is a diagram illustrating the procedure of the mass spectrometry method according to one embodiment. 
         FIG. 7  is a diagram illustrating the procedure of the mass spectrometry method according to one embodiment. 
     
    
    
     DESCRIPTION OF EMBODIMENTS 
     Hereinafter, preferred embodiments of the present disclosure will be described in detail, with reference to the drawings. Note that, in each of the drawings, the same reference numerals will be applied to the same portions or the corresponding portions, and the repeated description will be omitted. In addition, dimensions or dimensional ratios of each member (or part) illustrated in the drawings may be different from actual dimensions or dimensional ratios in order to make the description easy to understand. 
     [Configuration of Sample Support] 
       FIG. 1  is a plan view of a sample support  1  of one embodiment. As illustrated in  FIG. 1  and  FIG. 2 , the sample support  1  includes a substrate  2 , a frame (a frame body)  3 , and a conductive layer  4 . The sample support  1  is a sample support for sample ionization. The sample support  1 , for example, is used for ionizing a component of a sample that is a measurement target, at the time of performing mass spectrometry. 
     The substrate  2  includes a first surface  2   a  and a second surface  2   b  on a side opposite to the first surface  2   a.  A plurality of through holes  2   c  are formed on the substrate  2  uniformly (with a homogeneous distribution). Each of the through holes  2   c  extends in a thickness direction of the sample support  1  (that is, the substrate  2 ) (hereinafter, simply referred to as a “thickness direction”) and opens to the first surface  2   a  and the second surface  2   b.  The thickness direction is a direction perpendicular to the first surface  2   a  and the second surface  2   b.  The substrate  2 , for example, is formed of an insulating material in a rectangular plate shape. The length of one side of the substrate  2  when viewed in the thickness direction, for example, is approximately several cm to several tens of cm. The thickness of the substrate  2 , for example, is approximately 1 μm to 50 μm. In this embodiment, the thickness of the substrate  2  is approximately 5 μm. The substrate  2  is approximately transparent with respect to visible light. For example, the sample described above can be visually recognized via the substrate  2 . 
     The frame  3  is provided on the first surface  2   a  of the substrate  2 . Specifically, the frame  3  is fixed to the first surface  2   a  of the substrate  2  by an adhesive layer  5 . It is preferable to use an adhesive material having less emitted gas (for example, low-melting glass, a vacuum adhesive agent, or the like), as the material of the adhesive layer  5 . The frame  3  has a rectangular frame shape. The frame  3  is provided in a peripheral portion of the substrate  2 . The frame  3  includes a rectangular inner edge  3   a  and a rectangular outer edge  3   b.  The frame  3  surrounds an effective region (an ionization region) R when viewed in the thickness direction. The effective region R is a region that functions in order to move the component of the sample described below to the first surface  2   a  side in the substrate  2  and to ionize the component of the sample. 
     The frame  3  has approximately the same outer shape as that of the substrate  2  when viewed in the thickness direction. The length of one side of the frame  3  when viewed in the thickness direction (the length of one side of the outer edge  3   b ), for example, is approximately several cm to several tens of cm. The length of one side of the inner edge  3   a  (the effective region R) of the frame  3  when viewed in the thickness direction, for example, is approximately several cm to several tens of cm. The thickness of the frame  3 , for example, is less than or equal to 1 mm. The material of the frame  3 , for example, is a metal, a ceramic, and the like. According to such a frame  3 , the handling of the sample support  1  is facilitated, and the modification of the substrate  2  due to a temperature change or the like is suppressed. 
     A first marker  50  and a second marker  60  are provided on a surface  3   c  of the frame  3  on a side opposite to the substrate  2 . A plurality of first markers  50  are disposed along an X axis direction in a first portion  31  of the frame  3  extending along the X axis direction (a first direction). The first marker  50 , for example, is a plurality of numeric characters arranged along the X axis direction. Similarly, a plurality of second markers  60  are disposed along a Y axis direction in a second portion  32  of the frame  3  extending along the Y axis direction (a second direction orthogonal to the first direction). The second marker  60 , for example, is a plurality of numeric characters arranged along the Y axis direction. The first marker  50  and the second marker  60  configure a coordinate system for recognizing a position in the effective region R when viewed in the thickness direction. The first marker  50  and the second marker  60 , for example, are formed by making the surface  3   c  of the frame  3  concave and convex. 
     The conductive layer  4  is provided on the first surface  2   a  of the substrate  2 . Specifically, the conductive layer  4  is continuously (integrally) formed in a region corresponding to the inner edge  3   a  of the frame  3  (that is, a region corresponding to the effective region R) on the first surface  2   a  of the substrate  2 , an inner surface of the inner edge  3   a,  and the surface  3   c  of the frame  3 . In the effective region R, the conductive layer  4  is provided in a peripheral portion of the through hole  2   c  on the first surface  2   a.  That is, the conductive layer  4  covers a portion in the first surface  2   a  of the substrate  2 , on which the through hole  2   c  is not formed. That is, the conductive layer  4  is provided not to block the through hole  2   c.  In the effective region R, each of the through holes  2   c  is exposed to the inner edge  3   a.  The conductive layer  4  covers the first marker  50  and the second marker  60  on the surface  3   c  of the frame  3 . However, the first marker  50  and the second marker  60  are formed by making the surface  3   c  concave and convex, and thus, the visual contact and the recognition of the device are not hindered even in the case of being covered with the conductive layer  4 . 
     The conductive layer  4  is formed of a conductive material. Here, it is preferable that a metal having low affinity (reactivity) with respect to a sample and high conductivity is used as the material of the conductive layer  4 , from the following reasons. 
     For example, in a case where the conductive layer  4  is formed of a metal such as copper (Cu) having high affinity with respect to a sample such as protein, in a process of ionizing the sample described below, the sample is ionized in a state where Cu atoms are attached to sample molecules, and thus, there is a concern that a detection result is shifted in a mass spectrometry method described below as the Cu atoms are attached. Therefore, it is preferable that a metal having low affinity with respect to the sample is used as the material of the conductive layer  4 . 
     On the other hand, a metal having high conductivity easily and stably applies a constant voltage. For this reason, in a case where the conductive layer  4  is formed of the metal having high conductivity, it is possible to homogeneously apply a voltage to the first surface  2   a  of the substrate  2 . In addition, there is a tendency that the metal having high conductivity also has high thermal conductivity. For this reason, in a case where the conductive layer  4  is formed of the metal having high conductivity, it is possible to efficiently transfer the energy of an energy ray such as laser beam that is applied to the substrate  2  to the sample via the conductive layer  4 . Therefore, it is preferable that the metal having high conductivity is used as the material of the conductive layer  4 . 
     From the viewpoint described above, for example, it is preferable that gold (Au), platinum (Pt), and the like are used as the material of the conductive layer  4 . The conductive layer  4 , for example, is formed to have a thickness of approximately 1 nm to 350 nm by a plating method, an atomic layer deposition (ALD) method, an evaporation method, a sputtering method, and the like. In this embodiment, the thickness of the conductive layer  4  is approximately 10 nm. Note that, for example, chromium (Cr), nickel (Ni), titanium (Ti), and the like may be used as the material of the conductive layer  4 . 
       FIG. 3  is a diagram illustrating an enlarged image of the substrate  2  when viewed in the thickness direction. In  FIG. 3 , a black portion is the through hole  2   c,  and a white portion is a partition portion between the through holes  2   c.  As illustrated in  FIG. 3 , the plurality of through holes  2   c  having an approximately constant width are uniformly formed on the substrate  2 . The through hole  2   c,  for example, is approximately in a circle shape when viewed in the thickness direction. The width of the through hole  2   c,  for example, is approximately 1 nm to 700 nm. In this embodiment, the width of the through hole  2   c  is approximately 200 nm. The width of the through hole  2   c  indicates the diameter of the through hole  2   c  in a case where the through hole  2   c  is approximately in a circle shape when viewed in the thickness direction, and indicates the diameter (an effective diameter) of a virtual maximum cylinder falling into the through hole  2   c  in a case where the through hole  2   c  is not approximately in a circle shape. A pitch between the respective through holes  2   c,  for example, is approximately 1 nm to 1000 nm. In a case where the through hole  2   c  is approximately in a circle shape when viewed in the thickness direction, the pitch between the respective through holes  2   c  indicates a center-to-center distance of the respective circles, and in a case where the through hole  2   c  is not approximately in a circle shape, the pitch between the respective through holes  2   c  indicates a center axis-to-center axis distance of the virtual maximum cylinder falling into the through hole  2   c.  The width of the partition portion between the through holes  2   c  on the substrate  2 , for example is approximately 300 nm. 
     An opening rate of the through holes  2   c  (a ratio of all of the through holes  2   c  to the first surface  2   a  when viewed in the thickness direction) is practically 10% to 80%, and is particularly preferably 50% to 80%. The sizes of the plurality of through holes  2   c  may be uneven with each other, and the plurality of through holes  2   c  may be partially connected to each other. 
     The substrate  2 , for example, is an alumina porous film that is formed by performing anodic oxidation with respect to aluminum (Al). Specifically, an anodic oxidation treatment is performed with respect to an Al substrate, and a surface portion that is oxidized is peeled off from the Al substrate, and thus, the substrate  2  can be obtained. Note that, the substrate  2  may be formed by performing anodic oxidation with respect to a valve metal other than Al, such as tantalum (Ta), niobium (Nb), titanium (Ti), hafnium (Hf), zirconium (Zr), zinc (Zn), tungsten (W), bismuth (Bi), and antimony (Sb), or may be formed by performing anodic oxidation with respect to silicon (Si). 
       FIG. 4  is an enlarged view of the frame  3 . As illustrated in  FIG. 4 , the first marker  50  includes a marker  51  for visual contact and a marker  52  for a device. The marker  51  for visual contact is a marker to be read by the visual contact of a measurer. A plurality of markers  51  for visual contact are disposed along the X axis direction. The plurality of markers  51  for visual contact, for example, are arranged at regular intervals along the X axis direction. In this embodiment, as an example, each of the markers  51  for visual contact is a numeric character. A width w 1  of the marker  51  for visual contact can be recognized by the visual contact of the measurer. The width w 1  of the marker  51  for visual contact is greater than or equal to a predetermined value (for example, 1 mm). 
     The width w 1  of the marker  51  for visual contact, for example, is approximately 1 mm to 4 mm. A pitch w 2  between the markers  51  for visual contact adjacent to each other (that is, a distance between the centers of the markers  51  for visual contact adjacent to each other), for example, is approximately 2 mm to 10 mm. A length w 3  of a space between the markers  51  for visual contact adjacent to each other, for example, is approximately 1 mm to 9 mm. In this embodiment, as an example, the width w 1  of the marker  51  for visual contact is approximately 1 mm, the pitch w 2  between the markers  51  for visual contact is approximately 2 mm, and the length w 3  of the space between the markers  51  for visual contact is approximately 1 mm. The marker  51  for visual contact, for example, is provided by forming concavities and convexities on the surface  3   c  of the frame  3 , in accordance with a punch mark using stamping or laser. An engraved height (a depth) of the marker  51  for visual contact, for example, is approximately 0.1 mm to 0.9 mm. 
     The marker  52  for a device, for example, is a marker to be read by a camera  16  (refer to  FIG. 7 ) that is attached to a mass spectrometry device  10  (an ionization device) described below. In this embodiment, as an example, the marker  52  for a device is positioned inside the frame  3  from the marker  51  for visual contact. A plurality of markers  52  for a device are disposed along the X axis direction. The plurality of markers  52  for a device, for example, are arranged at regular intervals along the X axis direction. In this embodiment, as an example, each of the markers  52  for a device is a numeric character. 
     A width w 4  of the marker  52  for a device is less than the width w 1  of the marker  51  for visual contact. That is, the width w 4  of the marker  52  for a device is less than a predetermined value. The width w 4  of the marker  52  for a device, for example, is approximately 1 nm to 0.1 mm. A pitch w 5  between the markers  52  for a device adjacent to each other (that is, a distance between the centers of the markers  52  for a device adjacent to each other) is less than the pitch w 2  of the marker  51  for visual contact. The pitch w 5  between the markers  52  for a device, for example, is approximately 5 nm to 0.2 mm. A length w 6  of a space between the markers  52  for a device adjacent to each other, for example, is approximately 4 nm to 0.1 mm. In this embodiment, as an example, the width w 4  of the marker  52  for a device is approximately 40 μm, the pitch w 5  between the markers  52  for a device is approximately 110 μm, and the length w 6  of the space between the markers  52  for a device is approximately 70 μm. The marker  52  for a device, for example, is provided by forming concavities and convexities on the surface  3   c  of the frame  3 , in accordance with a punch mark (as an example, shallow engraving) using laser. An engraved height (a depth) of the marker  52  for a device, for example, is approximately 5 μm. Note that, as the punch mark using the laser, for example, a method such as deep engraving may be applied. The engraved height of the marker  52  for a device is different in accordance with each method. 
     Note that, a positional relationship between the plurality of markers  51  for visual contact and the plurality of markers  52  for a device, for example, is stored in a correspondence table that is prepared in advance. The measurer is capable of grasping that a marker “ 14 ” or “ 15 ” of the marker  52  for a device corresponds to a center position of a marker “ 1 ” of the marker  51  for visual contact in the X axis direction, or an intermediate position of markers “ 1 ” and “ 2 ” of the marker  51  for visual contact in the X axis direction corresponds to a marker “ 22 ” of the marker  52  for a device, with reference to such a correspondence table. Note that, the correspondence table may be saved on paper or the like, or may be saved in a storage device (a memory, a storage, or the like) of a computer as data. 
     As with the first marker  50 , the second marker  60  also includes a marker  61  for visual contact (refer  FIG. 1 ) and a marker  62  for a device (refer  FIG. 1 ). That is, the marker  61  for visual contact is a marker to be read by the visual contact of the measurer. A plurality of markers  61  for visual contact are disposed along the Y axis direction. The plurality of markers  61  for visual contact, for example, are arranged at regular intervals along the Y axis direction. In this embodiment, as an example, each of the markers  61  for visual contact is a numeric character. The width of the marker  61  for visual contact, a pitch between the adjacent markers  61  for visual contact, and the length of a space between the adjacent markers  61  for visual contact are identical to the width w 1  of the marker  51  for visual contact, the pitch w 2  between the adjacent markers  51  for visual contact, and the length w 3  of the space between the adjacent markers  51  for visual contact, described above. The marker  62  for a device, for example, is a marker to be read by the camera  16  that is attached to the mass spectrometry device  10 . In this embodiment, as an example, the marker  62  for a device is positioned inside the frame  3  from the marker  61  for visual contact. The plurality of markers  62  for a device are disposed along the Y axis direction. The plurality of markers  62  for a device, for example, are arranged at regular intervals along the Y axis direction. In this embodiment, as an example, each of the markers  62  for a device is a numeric character. The width of the marker  62  for a device, a pitch between the adjacent markers  62  for a device, and the length of a space between the adjacent markers  62  for a device are identical to the width w 4  of the marker  52  for a device, the pitch w 5  between the adjacent markers  52  for a device, and the length w 6  of the space between the adjacent markers  52  for a device, described above. 
     [Sample Ionization Method] 
     Next, a sample ionization method using the sample support  1  will be described with reference to  FIG. 5  to  FIG. 7 . Here, as an example, a laser desorption/ionization method using laser beam (an energy ray) (a part of a mass spectrometry method of a mass spectrometry device  10 ) will be described. In  FIG. 5  and  FIG. 7 , the through hole  2   c,  the conductive layer  4 , and the adhesive layer  5  in the sample support  1  are not illustrated. 
     First, as illustrated in (a) of  FIG. 5 , a sample S is prepared (a first step). Specifically, the sample S is mounted on a mounting surface  6   a  of a glass slide (a mounting portion)  6 . The glass slide  6  is a glass substrate on which a transparent conductive film such as an indium tin oxide (ITO) film is formed, and the surface of the transparent conductive film is the mounting surface  6   a.  Note that, not only the glass slide  6  but also a member that is capable of ensuring conductivity (for example, a substrate formed of a metal material such as stainless steel, or the like) can be used as the mounting portion. Here, the sample S, for example, is a biological sample (a hydrous sample). The sample S, for example, is a liver slice of a mouse, or the like. In order to smoothly move a component S 1  of the sample S (refer to (b) of  FIG. 6 ), a solution for decreasing the viscosity of the component S 1  (for example, an acetonitrile mixed liquid, acetone, or the like) may be added to the sample S. 
     Subsequently, as illustrated in (b) of  FIG. 5 , the sample support  1  described above is prepared (the first step). The sample support  1  may be prepared by being manufactured by a person who carries out the ionization method and the mass spectrometry method, or may be prepared by being acquired from a manufacturer, a seller, or the like of the sample support  1 . Subsequently, the sample support  1  is disposed on the sample S such that the second surface  2   b  faces the sample S (a second step). The sample support  1  is disposed on the sample S such that the second surface  2   b  is in contact with the sample S. 
     Subsequently, as illustrated in (c) of  FIG. 5 , the sample support  1  is fixed to the glass slide  6 . The sample support  1  is fixed to the glass slide  6  by a tape  7  having conductivity (for example, a carbon tape or the like). The tape  7  fixes the sample support  1  such that the first marker  50  and the second marker  60  are exposed. That is, the first marker  50  and the second marker  60  are not covered with the tape  7 . Alternatively, for example, the tape  7  is formed of a transparent material, and thus, in a case where the first marker  50  and the second marker  60  can be read by the camera  16  described below even when the first marker  50  and the second marker  60  are covered with the tape  7 , the first marker  50  and the second marker  60  may be covered with the tape  7 . The tape  7  may be a part of the sample support  1 , or may be prepared separately from the sample support  1 . In a case where the tape  7  is a part of the sample support  1  (that is, in a case where the sample support  1  includes the tape  7 ), for example, the tape  7  may be fixed in advance to the surface  3   c  side of the frame  3 . More specifically, the tape  7  may be fixed onto the conductive layer  4  that is formed on the surface  3   c  of the frame  3 . 
     The component S 1  of the sample S is moved toward the first surface  2   a  side of the substrate  2  from the second surface  2   b  side of the substrate  2  through the through hole  2   c  by a capillary action. The component S 1  that is moved to the first surface  2   a  side of the substrate  2  is accumulated on the first surface  2   a  side by a surface tension.  FIG. 6  is a plan view illustrating a state in which the sample support  1  is disposed on the sample S. As illustrated in  FIG. 6 , the component S 1  of the sample S is moved to the first surface  2   a  side in a region D 1  of the effective region R (an existence range of the sample S). Subsequently, the camera  16  scans the first marker  50  and the second marker  60  provided in the frame  3 , and thus, an irradiation range D 2  of laser beam L in the effective region R is recognized by a control unit  17  (refer to  FIG. 7 ) (a third step). 
     Specifically, first, the measurer determines the irradiation range D 2 , on the basis of the region D 1  and the marker  51  for visual contact and the marker  61  for visual contact. More specifically, in a case where the sample support  1  is disposed on the sample S, the measurer, for example, determines a region including the region D 1  as the irradiation range D 2 , by the visual contact. In this embodiment, as an example, the irradiation range D 2  has a rectangle shape surrounded by a pair of side portions extending in the X axis direction and a pair of side portions extending in the Y axis direction. Then, the measurer grasps coordinates (X1,Y1) and (X2,Y2) of a start point P 1  and an end point P 2  of the irradiation range D 2 , on the basis of the marker  51  for visual contact and the marker  61  for visual contact, respectively. The coordinates (X1,Y1) and (X2,Y2) are coordinates in a coordinate system configured of the marker  51  for visual contact and the marker  61  for visual contact. Here, as an example, (X1,Y1) is (7.9,1.1), and (X2,Y2) is (1.5,5.9). Note that, the coordinates that are grasped as described above are values presumed by the visual contact of the measurer. 
     Subsequently, as illustrated in  FIG. 7 , in a state where the sample S is disposed between the glass slide  6  and the sample support  1 , the glass slide  6 , the sample support  1 , and the sample S are mounted on a support portion  12  of the mass spectrometry device  10 . 
     The mass spectrometry device  10  includes the support portion  12 , a sample stage  18 , the camera  16  (a scanning unit), an irradiation unit  13 , a voltage application unit  14 , an ion detection unit  15 , and the control unit  17 . The sample S or the like that is a spectrometry target is mounted on the support portion  12 . The support portion  12  on which the sample S or the like is mounted is mounted on the sample stage  18 . The sample S or the like mounted on the support portion  12  is observed by the camera  16 . Here, the width of an observation range (a visual field) C (refer to  FIG. 4 ) of the camera  16 , for example, is approximately 1.5 mm. That is, the visual field of the camera  16  is smaller than the effective region R, and has at least a size capable of observing the markers  52  and  62  for a device. The irradiation unit  13  irradiates the first surface  2   a  of the sample support  1  with the energy ray such as the laser beam L. The voltage application unit  14  applies a voltage to the first surface  2   a  of the sample support  1 . The ion detection unit  15  detects ions of the sample S that is ionized. The control unit  17  controls the operations of the sample stage  18 , the camera  16 , the irradiation unit  13 , the voltage application unit  14 , and the ion detection unit  15 . The control unit  17 , for example, is a computer device including a processor (for example, a central processing unit [CPU]), a memory (for example, a read only memory [ROM], a random access memory [RAM], or the like), and the like. 
     Subsequently, the control unit  17  recognizes irradiation range D 2 , on the basis of the marker  52  for a device and the marker  62  for a device. For example, first, the control unit  17  conveys the sample support  1  to the sample stage  18  up to a position in which the marker  52  for a device provided in the frame  3  is imaged by the camera  16 . Specifically, the control unit  17  positions the sample support  1  such that a target marker  52  for a device falls within the observation range C of the camera  16  (refer to  FIG. 4 ), by moving the sample stage  18  such that the camera  16  scans the markers  52  for a device arrayed along the X axis direction. Here, the control unit  17  acquires in advance the marker  52  for a device corresponding to the coordinate X1 (here, 7.9) of the start point P 1  and the coordinate X2 (here, 1.5) of the end point P 2 , which are grasped by the visual contact of the measurer. Such acquisition of the marker  52  for a device, for example, can be attained by an input manipulation of the measurer with respect to a computer configuring the control unit  17 . Then, the control unit  17  recognizes (stores) the X coordinate of the start point P 1  of the irradiation range D 2 , on the basis of a position x 1  of the sample stage  18  in the X axis direction when the marker  52  for a device corresponding to the coordinate X1 of the start point P 1  is read by the camera  16 . That is, the control unit  17  recognizes the position x 1  as the X coordinate of the start point P 1 . Similarly, the control unit  17  recognizes (stores) the X coordinate of the end point P 2  of the irradiation range D 2 , on the basis of a position x 2  of the sample stage  18  in the X axis direction when the marker  52  for a device corresponding to the coordinate X2 of the end point P 2  is read by the camera  16 . That is, the control unit  17  recognizes the position x 2  as the X coordinate of the end point P 2 . 
     Similarly, the control unit  17  conveys the sample support  1  to the sample stage  18  up to a position in which the marker  62  for a device provided in the frame  3  is imaged by the camera  16 . Specifically, the control unit  17  positions the sample support  1  such that a target marker  62  for a device falls within the observation range C of the camera  16 , by moving the sample stage  18  such that the camera  16  scans the markers  62  for a device arrayed along the Y axis direction. Here, the control unit  17  acquires in advance the marker  62  for a device corresponding to the coordinate Y 1  (here, 1.1) of the start point P 1  and the coordinate Y2 (here, 5.9) of the end point P 2 , which are grasped by the visual contact of the measurer. Such acquisition of the marker  62  for a device can be attained by the same method as the acquisition of the marker  52  for a device described above. Then, the control unit  17  recognizes (stores) the Y coordinate of the start point P 1  of the irradiation range D 2 , on the basis of a position y 1  of the sample stage  18  in the Y axis direction when the marker  62  for a device corresponding to the coordinate Y1 of the start point P 1  is read by the camera  16 . That is, the control unit  17  recognizes the position y 1  as the Y coordinate of the start point P 1 . Similarly, the control unit  17  recognizes (stores) the Y coordinate of the end point P 2  of the irradiation range D 2 , on the basis of a position y 2  of the sample stage  18  in the Y axis direction when the marker  62  for a device corresponding to the coordinate Y2 of the end point P 2  is read by the camera  16 . That is, the control unit  17  recognizes the position y 2  as the Y coordinate of the end point P 2 . 
     As described above, the control unit  17  is capable of recognizing the positions (x 1 ,y 1 ) and (x 2 ,y 2 ) of the start point P 1  and the end point P 2 , respectively, by causing the camera  16  to scan the plurality of markers  52  for a device arrayed along the X axis direction and the plurality of markers  62  for a device arrayed along the Y axis direction. Accordingly, the control unit  17  recognizes the irradiation range D 2 . Note that, the coordinates (X1,Y1) and (X2,Y2) are the coordinates in the coordinate system configured of the marker  51  for visual contact and the marker  61  for visual contact of the frame  3 , whereas the positions (x 1 ,y 1 ) and (x 2 ,y 2 ) are a control coordinate system that is used by the control unit  17 . That is, the positions (x 1 ,y 1 ) and (x 2 ,y 2 ) are a position referred to when the irradiation unit  13  performs scanning Here, the coordinates (X1,Y1) of the start point P 1  and the coordinates (X2,Y2) of the end point P 2  correspond to the positions (x 1 ,y 1 ) and (x 2 ,y 2 ). From such a correspondence relationship, the coordinate system configured of the marker  51  for visual contact and the marker  61  for visual contact of the frame  3  and the control coordinate system can be exchanged each other. 
     Subsequently, a voltage is applied to the conductive layer  4  of the sample support  1  (refer to  FIG. 2 ) via the mounting surface  6   a  of the glass slide  6  and the tape  7  by the voltage application unit  14  (a fourth step). Subsequently, the control unit  17  operates the irradiation unit  13 , on the basis of irradiation range D 2  that is recognized by the control coordinate system (that is, a range that is specified by the positions (x 1 ,y 1 ) and (x 2 ,y 2 )). Specifically, the control unit  17  operates the irradiation unit  13  such that the first surface  2   a  in the irradiation range D 2  is irradiated with the laser beam L (the fourth step). Accordingly, the irradiation unit  13  scans the first surface  2   a  in the irradiation range D 2  with the laser beam L. 
     As an example, the control unit  17  moves the sample stage  18 , and controls an irradiation operation (an irradiation timing or the like) of the laser beam L by the irradiation unit  13 , as the control of the operation of the irradiation unit  13 . That is, the control unit  17  checks that the sample stage  18  is moved by a predetermined interval, and then, executes the irradiation of the laser beam L with respect to the irradiation unit  13 . Specifically, first, the control unit  17  moves the sample stage  18 , and thus, the irradiation position of the laser beam L by the irradiation unit  13  is adjusted to the position (x 1 ,y 1 ) corresponding to the start point P 1  of the irradiation range D 2 . Then, the laser beam L is applied to the position (x 1 ,y 1 ) of the start point P 1 . Subsequently, the control unit  17  moves the sample stage  18 , and thus, the irradiation position of the laser beam L by the irradiation unit  13  is adjusted to a position separated from the position (x 1 ,y 1 ) in the X axis direction by a predetermined interval (a laser irradiation interval set in advance), and the laser beam L is applied to the position. By repeating such a manipulation, the laser beam L is sequentially applied at each predetermined interval in the X axis direction. Then, in a case where the irradiation position of the laser beam L reaches an edge portion of the irradiation range D 2  (that is, a position (x 2 ,y 1 ) corresponding to coordinates (X3,Y3) of a turn-around point P 3  in  FIG. 6 ), the control unit  17  moves the sample stage  18 , and thus, the irradiation position of the laser beam L by the irradiation unit  13  is adjusted to a position separated from the immediately preceding irradiation position in the Y axis direction by a predetermined interval, and the laser beam L is applied to the position. Subsequently, the laser beam L is applied to a position separated from the immediately preceding irradiation position in the X axis direction by a predetermined interval. The laser beam L sequentially applied at each predetermined interval in the X axis direction. As described above, the laser beam L is scanned in a meander shape in the irradiation range D 2 , and then, reaches the position (x 2 ,y 2 ) of the end point P 2 . As described above, the first surface  2   a  in the irradiation range D 2  is scanned with the laser beam L. Note that, the scanning of the laser beam L with respect to the first surface  2   a  can be carried out by operating at least one of the sample stage  18  and the irradiation unit  13 . In a case where the irradiation unit  13  is operated, the control unit  17  controls the operation of the irradiation unit  13 , and controls both of the movement of the irradiation unit  13  and the irradiation operation of the laser beam L by the irradiation unit  13 . 
     As described above, the first surface  2   a  in the irradiation range D 2  is irradiated with the laser beam L while a voltage is applied to the conductive layer  4 , and thus, the component S 1  that is moved to the first surface  2   a  side through the through hole  2   c  in the irradiation range D 2  is ionized, and a sample ion S 2  (the component S 1  that is ionized) is emitted. Specifically, energy is transmitted from the conductive layer  4  absorbing the energy of the laser beam L to the component S 1  that is moved to the first surface  2   a  side of the substrate  2 , and the component S 1  obtaining the energy is gasified and obtains a charge, and thus, the sample ion S 2  is obtained. Each of the steps described above corresponds to the ionization method of the sample S, using the sample support  1  (here, as an example, a laser desorption/ionization method as a part of the mass spectrometry method). 
     The sample ion S 2  that is emitted is moved toward a ground electrode (not illustrated) that is provided between the sample support  1  and an ion detection unit  15  while being accelerated. That is, the sample ion S 2  is moved toward the ground electrode while being accelerated by a potential difference that occurs between the conductive layer  4  to which a voltage is applied and the ground electrode. Then, the sample ion S 2  is detected by the ion detection unit  15  (a fifth step). 
     A detection result of the sample ion S 2  by the ion detection unit  15  is associated with the irradiation position of the laser beam L. Specifically, the ion detection unit  15  detects the sample ion S 2  in each position in the irradiation range D 2  to which the laser beam L is applied, as described above. An identification number indicating each position (that is, the position to which the laser beam L is applied) (for example, the coordinates or the like in the control coordinate system, such as (x 1 ,y 1 ) described above) is applied to the data (the detection result) of the sample ion S 2  detected in each position in the irradiation range D 2 . Accordingly, a distribution image (MS mapping data) indicating a mass distribution of the sample S in the irradiation range D 2  is acquired. Further, it is possible to image a two-dimensional distribution of molecules configuring the sample S. Note that, here, the mass spectrometry device  10  is a mass spectrometry device using a time-of-flight mass spectrometry (TOF-MS) method. 
     Subsequently, in a state where the sample support  1  is disposed on the sample S, an optical image of the sample S and the sample support  1  is acquired (a sixth step). Here, the acquired optical image includes at least of the region D 1  in the effective region R, and the first marker  50  and the second marker  60 . Subsequently, the optical image is superimposed on the distribution image such that the irradiation range D 2  in the optical image overlaps with the distribution image of the sample S, on the basis of the marker  51  for visual contact and the marker  61  for visual contact in the optical image (a seventh step). Specifically, the position (x 1 ,y 1 ) of the start point P 1  and the position (x 2 ,y 2 ) of the end point P 2  in the distribution image are superimposed on the coordinates (X1,Y1) of the start point P 1  and the coordinates (X2,Y2) of the end point P 2  in the optical image, respectively. Accordingly, the optical image of the sample S and the distribution image are synthesized. Note that, the substrate  2  is approximately transparent with respect to visible light, and thus, even in a case where the sample support  1  is disposed on the sample S, it is possible to acquire the optical image of the sample S. Each of the steps described above corresponds to the mass spectrometry method using the sample support  1 . 
     After the observation (optical observation) of the optical image of the sample S and the distribution image of the sample S that are synthesized together, detailed screening of a liquid chromatography-mass spectrometry (LC-MS) method may be further performed with respect to a portion having specific data in the sample S. In such a case, for example, an identification number such as the information of the irradiation position is applied to the specific data, and thus, it is possible to specify the irradiation position and the coordinates in the sample support  1 , on the basis of the identification number. Accordingly, it is possible to supply a sample of the sample S in the coordinates to LC-MS. 
     As described above, in the sample support  1 , the plurality of through holes  2   c  opening to the first surface  2   a  and the second surface  2   b  on a side opposite to the first surface  2   a  are formed on the substrate  2 . For this reason, in a case where the sample support  1  is disposed on the sample S such that the second surface  2   b  of the substrate  2  faces the sample S, it is possible to move the component  51  of the sample S toward the first surface  2   a  side from the second surface  2   b  side through the through hole  2   c  by using a capillary action. Further, in a case where the first surface  2   a  is irradiated with the laser beam, the energy is transmitted to the component  51  of the sample S that is moved to the first surface  2   a  side via the conductive layer  4 , and thus, it is possible to ionize the component S 1  of the sample S. In addition, the sample support  1  includes the frame  3  provided in the peripheral portion of the substrate  2 . For this reason, it is possible to improve the handleability of the sample support  1  by the frame  3 . In addition, the frame  3  surrounds the effective region R in which the sample S is ionized when viewed in the thickness direction of the substrate  2 , and the first marker  50  and the second marker  60  for recognizing the position in the effective region R are provided in the frame  3 . Accordingly, the following effects are obtained. 
     That is, in this embodiment, in a case where the visual field of the camera  16  is narrow, and the irradiation range D 2  is specified by observing the effective region R (the entire sample S), it is difficult to determine the irradiation position of the laser beam L. According to the sample support  1 , it is possible for the mass spectrometry device  10  to recognize the irradiation range D 2  of the laser beam L, by causing the camera  16  to perform scanning and by reading the first marker  50  and the second marker  60  provided in the frame  3 . Accordingly, according to the sample support  1 , it is possible to easily recognize the irradiation range D 2  of the laser beam L. In particular, in a mass spectrometry device (an MALDI-MS device) that is used in existing MALDI, the number of pixels, the visual field, or the like of the camera that is attached to the device is optimized to be suitable for the observation of matrix crystals in the sample, and thus, there is a problem that the mass spectrometry device is not suitable for the determination of the irradiation position of the laser beam L and the observation of the entire sample for imaging. It is possible to solve the problem of the MALDI-MS device described above by applying the mass spectrometry device  10  described above to such an MALDI-MS device. 
     The width of the through hole  2   c  is 1 nm to 700 nm, and the thickness of the substrate  2  is 1 μm to 50 μm. Accordingly, it is possible to suitably attain the movement of the component S 1  of the sample S by the capillary action described above. 
     The plurality of first markers  50  disposed along the X axis direction are provided in the first portion  31  extending along the X axis direction of the frame  3 , and the plurality of second markers  60  disposed along the Y axis direction are provided in the second portion  32  extending along the Y axis direction orthogonal to the X axis direction of the frame  3 . Accordingly, it is possible to recognize the position in the X axis direction by the first marker  50  and to recognize the position in the Y axis direction by the second marker  60 . Accordingly, it is possible to easily grasp two-dimensional coordinates of the irradiation range D 2  of the laser beam L (for example, the position of the start point P 1 , the position of the end point P 2 , or the like). 
     The first marker  50  and the second marker  60  are a numeric character. Accordingly, it is possible to attain a marker suitable for visual contact and/or for reading a device. 
     The first marker  50  includes the marker  51  for visual contact having the width w 1  of greater than or equal to the predetermined value and the marker  52  for a device having the width w 4  of less than the predetermined value, and the second marker  60  includes the marker  61  for visual contact having a width of greater than or equal to a predetermined value and a marker  62  for a device having a width of less than the predetermined value. Accordingly, the markers  51  and  61  for visual contact are read by the visual contact of the measurer, and thus, it is possible to determine in advance the irradiation range D 2 . Further, the markers  52  and  62  for a device corresponding to the irradiation range D 2  determined by the measurer are read by the camera  16 , and thus, it is possible for the mass spectrometry device  10  to recognize the irradiation range D 2  of the laser beam L. 
     In addition, in the ionization method of the sample S described above, the plurality of through holes  2   c  opening to the first surface  2   a  and the second surface  2   b  on a side opposite to the first surface  2   a  are formed on the substrate  2 . In a case where the sample support  1  is disposed on the sample S such that the second surface  2   b  of the substrate  2  faces the sample S, the component  51  of the sample S is moved toward the first surface  2   a  side from the second surface  2   b  side through the through hole  2   c  by the capillary action. Further, in a case where the first surface  2   a  is irradiated with the laser beam L while a voltage is applied to the conductive layer  4 , the energy is transmitted to the component  51  of the sample S that is moved to the first surface  2   a  side. Accordingly, the component  51  of the sample S is ionized. In addition, it is possible for the mass spectrometry device  10  to easily recognize the irradiation range D 2  of the laser beam L by scanning the first marker  50  and the second marker  60  provided in the frame  3 . 
     In the ionization method, the first marker  50  includes the marker  51  for visual contact having the width w 1  of greater than or equal to the predetermined value and the marker  52  for a device having the width w 4  of less than the predetermined value, and the second marker  60  includes the marker  61  for visual contact having the width of greater than or equal to the predetermined value and the marker  62  for a device having the width of less than the predetermined value. Then, in the third step, the measurer determines the irradiation range D 2 , on the basis of the region D 1  in the effective region R, and the markers  51  and  61  for visual contact, and the control unit  17  recognizes the irradiation range D 2 , on the basis of the position of the sample stage  18  when the markers  52  and  62  for a device corresponding to the irradiation range D 2  determined by the measurer are read by the camera  16 . Accordingly, it is possible to accurately attain both of the determination of the irradiation range D 2  by the visual contact of the measurer and the recognition of the irradiation range D 2  by a mechanical manipulation (marker scanning) of the mass spectrometry device  10 , by the first marker  50  and the second marker  60  provided in the frame  3 . 
     As described above, according to the mass spectrometry method described above, it is possible to accurately superimpose the optical image of the sample S on the distribution image, on the basis of the first marker  50  and the second marker  60  provided in the frame  3  of the sample support  1 . As a result thereof, it is possible to visualize the mass distribution in each position of the sample S. 
     [Modification Example] 
     As described above, the embodiment of the present disclosure has been described, but the present disclosure is not limited to the embodiment described above, and the present disclosure can be variously modified within a range not departing from the gist thereof. 
     The substrate  2  may have conductivity, and in the mass spectrometry method, the first surface  2   a  may be irradiated with the laser beam L while a voltage is applied to the substrate  2 . In a case where the substrate  2  has conductivity, it is possible to omit the conductive layer  4  in the sample support  1  and to obtain the same effects as those in the case of using the sample support  1  including the conductive layer  4  described above. Note that, irradiating the first surface  2   a  of the substrate  2  with the laser beam L indicates that the conductive layer  4  is irradiated with the laser beam L in a case where the sample support  1  includes the conductive layer  4 , and indicates that the first surface  2   a  of the substrate  2  is irradiated with the laser beam L in a case where the substrate  2  has conductivity. 
     An example has been described in which the first marker  50  and the second marker  60  are a numeric character, but the first marker  50  and the second marker  60  may be various markers. The first marker  50  and the second marker  60 , for example, may be at least one selected from a numeric character, a signal, and a letter. Even in this case, it is possible to attain a marker suitable for visual contact and/or for reading a device. Each of the marker  51  for visual contact, the marker  52  for a device, the marker  61  for visual contact, and the marker  62  for a device, for example, may be at least one selected from a numeric character, a signal, and a letter. In addition, such markers may include auxiliary information such as a graduation line. 
     An example has been described in which the first marker  50  and the second marker  60  are formed by making the surface  3   c  of the frame  3  concave and convex, but the first marker  50  and the second marker  60  may not be formed by making the surface  3   c  of the frame  3  concave and convex. The first marker  50  and the second marker  60 , for example, may be formed by a print according to printing such as nanoprinting, photolithography using extreme ultraviolet (EUV) lithographic exposure, write according to a paint, black (oxidization) that is an example of a punch mark using a laser, foamed marking using a laser, or the like. In a case where the first marker  50  and the second marker  60  are formed by the foamed marking using the laser, the material of the frame  3  is a resin. The foamed marking using the laser is a method for foaming the resin by a laser beam. According to such a method, light is diffusely reflected in a foamed portion, and as a result thereof, the visibility of the foamed portion increases. Note that, in a case where the first marker  50  and the second marker  60  are not formed by making the surface  3   c  of the frame  3  concave and convex and the visibility is inhibited at the time of being covered with the conductive layer  4 , the conductive layer  4  may not be formed in a region on the surface  3   c  of the frame  3 , in which the first marker  50  and the second marker  60  are formed. 
     The frame  3  may include only one of the first marker  50  and the second marker  60 . In this case, it is possible for the mass spectrometry device  10  to recognize a range in at least one direction of the irradiation range D 2  in the X axis direction and the Y axis direction. In addition, the first marker  50  may be provided in both of the first portions  31  facing each other in the frame  3 . Similarly, the second marker  60  may be provided in both of the second portions  32  facing each other in the frame  3 . 
     The first marker  50  may include only one of the marker  51  for visual contact and the marker  52  for a device. In this case, it is preferable that the width of the first marker  50  can be read by any of the visual contact of the measurer and the camera  16  that is attached to the mass spectrometry device  10 . Similarly, the second marker  60  may include only one of the marker  61  for visual contact and the marker  62  for a device. In this case, it is preferable that the width of the second marker  60  can be read by any of the visual contact of the measurer and the camera  16  that is attached to the mass spectrometry device  10 . 
     An example has been described in which the determination of the irradiation range D 2  by the measurer is performed before the glass slide  6 , the sample support  1 , and the sample S are mounted on the support portion  12  of the mass spectrometry device  10 , but such determination of the irradiation range D 2  by the measurer may be performed after the glass slide  6 , the sample support  1 , and the sample S are mounted on the support portion  12  of the mass spectrometry device  10 . 
     An example has been described in which the optical image of the sample S and the sample support  1  is acquired in a state where the sample support  1  is disposed on the sample S (the sixth step) is performed after the distribution image is acquired (the fifth step), but such acquisition of the optical image may be performed at any time after the sample support  1  is disposed on the sample S (the second step). For example, such an optical image may be acquired before the glass slide  6 , the sample support  1 , and the sample S are mounted on the support portion  12  of the mass spectrometry device  10 . 
     In the third step, as with the start point P 1  and the end point P 2 , the coordinates (X3,Y3) of the turn-around point P 3  (refer to  FIG. 6 ) may be grasped, and the coordinates of the turn-around point P 3  of the irradiation range D 2  may be recognized (stored), on the basis of the position (x 2 ,y 1 ) of the sample stage  18  when the markers  52  and  62  for a device corresponding to the coordinates (X3,Y3) of the turn-around point P 3  are read. In a case where there is the information of the turn-around point P 3 , in addition to the start point P 1  and the end point P 2 , it is possible to more accurately superimpose the optical image of the sample S on the distribution image. For example, in a case where the optical image is superimposed on the distribution image, and then, it is necessary to match the direction such as an up-down direction and a right-left direction, it is possible to suitably superimpose the optical image on the distribution image, on the basis of the information of three points not on a straight line (the start point P 1 , the end point P 2 , and the turn-around point P 3 ). In addition, in a case where the irradiation range D 2  is not in a rectangle shape (for example, the irradiation range D 2  is in a parallelogram shape in which one side portion of four side portions intersects with the Y axis direction), the information of the turn-around point P 3  may be read in a case where three or more reference points (coordinates) are necessary for specifying the irradiation range D 2 . 
     An example has been described in which the control unit  17  moves the sample stage  18  such that the camera  16  scans the markers  52  and  62  for a device, but the scanning of the markers  52  and  62  for a device by the camera  16  can be carried out by operating at least one of the sample stage  18  and the camera  16 . In a case where the camera  16  is operated, the control unit  17  recognizes the irradiation range D 2 , on the basis of the position of the camera  16  when the markers  52  and  62  for a device corresponding to the irradiation range D 2  determined by the measurer is read by the camera  16 . Accordingly, as with a case where the sample stage  18  is operated, it is possible to accurately attain both of the determination of the irradiation range D 2  by the visual contact of the measurer and the recognition of the irradiation range D 2  by the mechanical manipulation (the marker scanning) of the mass spectrometry device  10 , by the first marker  50  and the second marker  60  provided in the frame  3 . 
     REFERENCE SIGNS LIST 
       1 : sample support,  2 : substrate,  2   a:  first surface,  2   b:  second surface,  2   c:  through hole,  3 : frame (frame body),  4 : conductive layer,  10 : mass spectrometry device,  13 : irradiation unit,  16 : camera (scanning unit),  17 : control unit,  31 : first portion,  32 : second portion,  50 : first marker,  51 ,  61 : marker for visual contact,  52 ,  62 : marker for device,  60 : second marker, D 1 : region (existence range), D 2 : irradiation range, L: laser beam (energy ray), R: effective region (ionization region), S: sample, S 1 : component, S 2 : sample ion, w 1 , w 4 : width.