JP6787682B2 - Sample loading plate and its manufacturing method - Google Patents

Description

The present invention relates to a sample loading plate for loading a sample and a method for producing the same.

A matrix-assisted laser desorption ionization (MALDI = Matrix Assisted Laser Desorption / Ionization) method is known as one of mass spectrometric ionization methods capable of rapidly and accurately diagnosing pathogenic bacteria and bacteria.

The MALDI method uses a substance (matrix) that easily absorbs laser light and easily ionizes it in order to analyze an object to be analyzed (hereinafter referred to as a sample) that is difficult to absorb laser light or is easily damaged by laser light. This is a method in which a sample is mixed in advance and the sample is ionized by irradiating the sample with a laser beam.

In a mass spectrometer using the MALDI method, generally, a sample to be analyzed and a matrix are mixed in advance and liquefied with a solvent (hereinafter referred to as a sample, which is liquid at the time of dropping, but a sample obtained by drying and crystallizing the sample A metal plate called a target plate (hereinafter referred to as a sample loading plate) on which a sample is loaded is placed in the apparatus, and the sample loaded on the sample loading plate is irradiated with laser light for a predetermined time. Desorb and ionize the sample. At this time, a voltage is applied to the metal sample loading plate, and an electric field is applied to the desorbed and ionized sample, so that the desorbed and ionized sample can easily fly toward the acceleration electrode.

The sample loading plate is provided with a plurality of sample loading areas (hereinafter referred to as sample loading spots) for loading samples, and a plurality of samples to be measured are dropped onto a predetermined sample loading spot and dried (crystallized). A plurality of samples are irradiated with a laser by arranging the sample in the mass spectrometer in a state of being in the state and moving the sample loading plate.

In the MALDI analysis method, such crystals are deposited as uniformly as possible in the sample loading spot, the object to be analyzed is appropriately desorbed and ionized, and an electric field is effectively applied to the sample from the voltage applied to the sample loading plate. Acceleration is important and many proposals have been made for these analytical techniques.

Regarding the improvement of sample crystallization or ionization in the sample loading spot, for example, the proposal shown in Patent Document 1 is that the sample loading spot has a central portion having an electrically conductive surface and a margin (peripheral) portion consisting of a hydrophobic mask. The sample dropped onto the sample loading spot is crystallized and deposited in a ring shape in the hydrophobic margin portion by the halo effect. The crystal ring formed in the margin portion is efficiently irradiated with laser light to perform ionization.

Further, in the proposal shown in Patent Document 2, a conductive interference layer is provided on an insulating substrate so as to exhibit a color different from that of the substrate, a hydrophobic film is formed on the surface, and a groove for forming a sample loading spot is further formed. Is provided to expose the substrate, and the dropped sample is retained in the sample loading spot (hereinafter referred to as an anchor effect) to be crystallized and ionized.

Special Table 2006-525525 WO2015 / 019861 US2002 / 0045270

However, in the prior art shown in Patent Document 1, although the crystal ring of the sample formed in the margin portion of the sample loading spot is irradiated with a laser beam to perform efficient measurement, the center having electrical conductivity is provided. Since the margin part is an insulating film with respect to the part, the conductivity is not sufficient, and the electric field is not effectively applied to the sample from the voltage applied to the sample loading plate for the deposited crystal ring, and the sample is charged up. However, there is a problem that proper ionization is hindered.

Further, the conventional technique shown in Patent Document 2 has an anchor effect of keeping the sample in the spot due to the effect of the groove, and the visibility of the sample is good because the color of the substrate is different from the sample, but the dropped sample is the sample. It is difficult to get wet and spread over the entire loading spot, and it is easy to get wet like a donut along the groove of the spot. As a result, the density of the sample in the center of the sample loading spot used for the analysis becomes low, which may lead to a decrease in the analysis sensitivity.

Further, in the prior art shown in Patent Document 3, a substrate or a metal surface formed on the substrate is exposed as a hydrophilic anchor that holds the sample in the center of the spot, and affinity adsorption is performed on the surface of the ring-shaped region surrounding the anchor. A sample support substrate having a structure in which a body is placed and a hydrophobic material such as Teflon (registered trademark) is placed so that the outer surface thereof becomes hydrophobic is disclosed. This configuration is an excellent configuration that enables the sample to be effectively concentrated on the anchor at the center of the spot and dried, but at the manufacturing stage, materials such as affinity adsorbents and hydrophobic materials are placed separately. There is a problem that the process is complicated because the process is required.

An object of the present invention is to solve the above problems, increase the anchoring effect of the sample at the time of loading the sample at the sample loading spot, and enable the sample to be concentrated and dried and crystallized at the center and near the center of the sample loading spot. It is to provide a sample loading plate and the manufacturing method thereof.

In order to achieve the above object, the configuration of the present invention is as follows.
A sample loading plate used for mass spectrometry of the MALDI method and having at least one sample loading spot for loading a sample on a substrate, the sample loading plate is located near the inner center of the edge of the sample loading spot. A hydrophilic surface is formed, a hydrophobic surface is formed at the margin, and the intermediate region between the vicinity of the inner center and the margin becomes less hydrophilic from the center of the sample loading spot toward the outside. It is characterized in that the surface is formed so as to change to hydrophobicity.

The intermediate region is characterized in that a hydrophilic surface and a hydrophobic surface are partially formed, respectively, and the area ratio of the hydrophobic surface increases from the center of the sample loading spot toward the outside.

The substrate is a resin, and the hydrophobic surface formed in the peripheral portion and the intermediate region is characterized in that the substrate is exposed.

The hydrophilic surface formed in the vicinity of the inner center and in the intermediate region is characterized in that the optical multilayer film formed on the substrate is exposed.

The substrate is ceramics or metal, and the hydrophilic surfaces formed in the vicinity of the inner center and the intermediate region are characterized in that the substrate is exposed.

A method for manufacturing a sample loading plate that is used for mass analysis of the MALDI method and has at least one sample loading spot for loading a sample on a substrate, and is a step of making the substrate surface a hydrophilic surface and a hydrophilic surface. In the process of forming a hydrophobic film on the surface to make it a hydrophobic surface, an opening is formed in the part near the center inside from the edge of the sample loading spot, and an opening is formed in the part with respect to the edge. An opening is formed in the portion of the intermediate region between the vicinity of the inner center and the edge portion so that the proportion of the opening decreases from the center of the sample loading spot toward the outside. It is characterized by having a step of placing the mask on the surface of the sample loading plate, a step of removing the hydrophobic film on the surface of the sample loading spot using the mask, and a step of removing the mask.

The mask has a plurality of concentric openings with respect to the center of the sample loading spot, and is open at the portion near the inner center, and there is no opening at the portion with respect to the edge portion, and the portion with respect to the intermediate region. The feature is that the ratio of the opening area gradually decreases from the center of the sample loading spot toward the outside.

The mask is characterized by having a plurality of linear openings extending radially from the center of the sample loading spot and having a constant width or a width decreasing from the center to the outside.

Another method of manufacturing a sample loading plate, which is used for mass spectrometry of the MALDI method and has at least one sample loading spot for loading a sample on a substrate, is a step of making the substrate surface a hydrophobic surface and a hydrophobic surface. In the process of forming a hydrophilic film on the sexual surface to make it a hydrophilic surface, and in the part of the sample loading spot near the inner center from the edge part, no opening is formed, and the part with respect to the edge part An opening is formed, and the proportion of the opening increases from the center of the sample loading spot toward the outside in the portion with respect to the intermediate region between the vicinity of the inner center and the edge portion. It is characterized by having a step of placing a mask having an opening formed on the surface of the sample loading plate, a step of removing the hydrophilic film on the surface of the sample loading spot using the mask, and a step of removing the mask. To do.

In the above alternative method, the mask has a plurality of concentric openings with respect to the center of the sample loading spot, has no opening in the portion near the inner center, and has an opening in the portion with respect to the edge portion. It is characterized in that the ratio of the opening area to the intermediate region gradually increases from the center of the sample loading spot toward the outside.

Similarly, in the above alternative method, the mask does not open at a site for a plurality of linear regions extending radially from the center of the sample loading spot and having a constant width or a width that decreases outward from the center. It is characterized by having a part.

According to the present invention, when the sample is loaded on the sample loading spot, the sample is wetted by the hydrophilic surface formed near the inner center of the sample loading spot and the hydrophilic surface formed in a part of the intermediate region. As a result, the sample can be reliably trapped by the sum of the areas near the inner center and the intermediate region of the sample loading spot, and a stronger anchor effect can be exhibited than when trapping only near the inner center. .. Then, as the sample dries and the volume decreases, the hydrophilic surface is attracted to the vicinity of the inner center where it is most exposed, and many samples can be kept in a dried and crystallized state near the inner center. It becomes. In manufacturing a sample loading plate that produces such a result, it is possible to easily manufacture the sample loading plate by removing the film on the outermost surface of the sample loading plate using one mask.

Sample loading plate according to Example 1 of the present invention The figure explaining the membrane structure of the sample loading plate in Example 1 of this invention. Partial sectional view explaining a state in which a sample is loaded on the sample loading spot in Example 1 of the present invention. The process drawing explaining the manufacturing method which concerns on the sample loading plate of Example 1 of this invention. Top view of the mask according to the manufacturing method of Example 1 of the present invention. Sample loading plate according to Example 2 of the present invention The figure explaining the membrane structure of the sample loading plate in Example 2 of this invention. Partial sectional view explaining the state which the sample was loaded on the sample loading spot in Example 2 of this invention. The process drawing explaining the manufacturing method which concerns on the sample loading plate of Example 2 of this invention. Top view of the mask according to the manufacturing method of Example 2 of the present invention. Sectional drawing schematically explaining the coloring principle by interference of an optical multilayer film Schematic diagram for explaining the operation of the mass spectrometer Enlarged view of part H of the sample loading spot in another embodiment of the present invention

Hereinafter, embodiments of the present invention will be described with reference to FIGS. 1 to 13.

The sample loading plate is mounted on a mass spectrometer (see FIG. 12 described later) by the MALDI method, and is used for loading a sample on a sample loading spot and analyzing the mass. The mode for carrying out the invention shown below exemplifies a sample loading plate and a method for manufacturing the sample loading plate for embodying the idea of the present invention, and the present invention specifies the method and configuration described below. It's not a thing. In particular, the manufacturing method and the shape, material, relative arrangement, etc. of the members described in the embodiments are not limited to the scope of the present invention unless otherwise specified. In addition, the size and shape of the members shown in each drawing, the positional relationship, and the film layer to be formed may be exaggerated for the sake of clarity.

[Summary Description of Sample Loading Plate of Embodiment: FIG. 1]
First, the configuration of the sample loading plate according to the embodiment of the present invention will be described with reference to FIG. FIG. 1 is a diagram relating to one of the two examples described in the present embodiment, but first, it is used to explain the schematic configuration of the present invention.

FIG. 1A is a plan view of the sample loading plate as viewed from the surface side on which the sample is loaded. The substrate 1 of the sample loading plate 100 is a substantially rectangular flat plate having an outer shape of about 50 mm × 40 mm, and the sample loading plate 100 is provided with a notch such as a lower side for positioning, for example. Further, the flatness of the sample loading plate 100 has an accuracy of 30 μm or less. The outer shape, thickness, etc. are not particularly limited as long as they meet the specifications of the mass spectrometer. The sample loading plate may be surface-finished by a wrapping step or a polishing step in order to ensure flatness.

The sample loading plate 100 has a column address mark 30 (for example, 1 to 9, X to Z) indicating the position of each sample loading spot 10, a row address mark 40 (for example, A to H), and a serial number 50 for managing the sample loading plate. , Bar code 60 and the like can be formed. These address marks, serial numbers, barcodes, etc. are not limited to these, and may be added or deleted as necessary. The method for forming the address mark, serial number, and barcode is not particularly limited, but a processing method by laser marking is preferable.

The sample loading spot 10 is an enlarged plan view shown in FIG. 1B, and is composed of an inner center vicinity 21 surrounded by a broken line and an intermediate region 24 surrounding the inner center vicinity 21. The inner center vicinity 21 is a hydrophilic surface, and the intermediate region 24 is surface-formed so that the ratio of the hydrophobic surface increases as the hydrophilic surface and the hydrophobic surface move from the center to the outside. The forming method will be described later. As described above, the sample loading spot 10 is defined as a region including the inner center vicinity 21 and the intermediate region 24. The outer peripheral portion 22 on the outer side of the intermediate region 24 has a structure in which a hydrophobic surface is formed, and the outer peripheral portion 22 of the adjacent sample loading spots and the loaded samples are mutually loaded. Separated enough to avoid mixing and contamination.

Next, two examples will be described with respect to the film and substrate cross-sectional structure of the sample loading plate 100 and the manufacturing method.

The first embodiment will be described with reference to FIGS. 1 to 5. FIG. 1B is a cross-sectional view taken along the cutting line I-I'passing through the center of the sample loading spot 10 shown in FIG. 1A, and is a plan view at the top in order to make it easy to understand the planar positional relationship. An enlarged view of is added. Here, the first metal film 2M is first formed on one side surface of the substrate 1. Next, the optical multilayer film 2A is formed by laminating on the first metal film 2M. The optical multilayer film 2A is made of a dielectric film or a second metal film, and the type and number of layers are not particularly limited, and are formed in the order of, for example, 2d, 2c, 2b, and 2a. The optical multilayer film 2A is hydrophilic and is defined as a first hydrophilic film. Further, the hydrophobic film 12 is formed on the optical multilayer film. The first metal film 2M and the optical multilayer film 2A are formed by a film forming method such as vacuum deposition or sputtering. The hydrophobic film 12 is also formed by a film forming method such as vacuum vapor deposition or sputtering, but it can also be formed by a method such as dip coating in which the film is immersed in a liquid and slowly pulled up to form a film.

As shown in FIG. 1B, the hydrophobic film 12 is peeled off in the vicinity of the inner center 21 in a plane, and the hydrophilic optical multilayer film 2A is exposed. In the intermediate region 24, the optical multilayer film 2A is partially exposed, and the hydrophobic film is partially peeled off so that the proportion of the hydrophobic film 12 increases toward the outside from the center of the sample mounting spot 10 to form a pattern. Is forming. The outer peripheral portion 22 outside the intermediate region 24 is covered with the hydrophobic film 12. Therefore, the hydrophobicity is gradually increased from the center of the sample loading spot to the outside.

FIG. 2 shows the material and thickness of each layer as the cross-sectional structure of the sample loading plate 100 in Example 1. Aluminium (Al ₂ O ₃ ) is used for the substrate 1. The first metal film 2M laminated on the substrate 1 uses Ni as a material and has a film thickness of about 300 nm (1 nm = 0.000001 mm). Next, the first layer 2d constituting the optical multilayer film 2A uses Al ₂ O _{3 and} has a film thickness of about 80 nm. Ti is used for the second layer 2c, and the film thickness is about 10 nm. The third layer 2b uses SiO _{2 and} has a film thickness of 90 nm. The fourth layer 2a uses Ti and has a film thickness of about 10 nm. With such a configuration, the surface of the sample loading plate 100 can be dark blue in the visible light wavelength region. A hydrophobic film 12 composed of C (carbon), F (fluorine), Si (silicon) and the like is formed on the optical multilayer film 2A. The thickness of the hydrophobic film 12 is, for example, 2 to 2. Since it is as thin as about 3 nm, it has little effect on the conductivity and color of the inner surface of the sample loading spot 10.

As described above, by preferably combining the first metal film laminated on the substrate 1 and the optical multilayer film 2A, arbitrary reflection characteristics (coloring) utilizing optical interference can be obtained.

Further, when the substrate 1 is exposed by the surface color of the sample loading plate and the groove formed in the vicinity of the inner center 21 by using a white material such as alumina for the substrate 1, the exposed substrate color becomes white and the optical multilayer film is formed. The contrast with 2A becomes even more remarkable, and the visibility of the sample loading spot 10 becomes better. Further, since the crystal of the sample is white, the surface of the sample loading plate can be color-identified from the optical multilayer film 2A, and the presence or absence of loading can be confirmed after crystallization. Further, if the substrate 1 is alumina, the grooves have a hydrophilic surface and can be more effective for the purpose of trapping the sample.

[Explanation of Manufacturing Method of Sample Loading Plate 110: FIGS. 4 and 5]
Next, the method of manufacturing the sample loading plate 100 of Example 1 will be described with reference to FIGS. 4 and 5. FIG. 4 is a process diagram showing a method of manufacturing the sample loading plate 100 according to the first embodiment. FIG. 5 is an example of a pattern of the mask 15 used in the manufacturing process.

[Explanation of manufacturing method: Fig. 4]
In FIG. 4, the main steps of 310 to 370 will be illustrated and described with respect to the method of manufacturing the sample loading plate 110. Unless otherwise specified in each step, it is natural to perform general operations such as transfer, inspection, cleaning, drying, and annealing necessary for each step, and the description thereof will be omitted.

[Substrate acceptance process: 310]
First, in the substrate receiving step 310, the flatness and surface roughness of the substrate 1 are inspected to confirm that the substrate 1 has a predetermined flatness and surface roughness.

[Substrate surface processing process (enlarged view): 320]
Next, in the substrate surface processing step 320, the substrate 1 is subjected to a lapping process or a polishing process to finish the substrate 1 to a predetermined substrate thickness, surface roughness, and flatness. The main inspection items in this step are the surface roughness and flatness of the substrate.

[First metal film forming step (enlarged view): 330]
Next, in the first metal film forming step 330, the first metal film 2M is formed. For example, Ni is formed to a thickness of 300 nm by using a film forming method such as vacuum deposition or sputtering. At this time, the irradiation direction of the film-formed particles is preferably vertical in order to make the film as uniform as possible (see the broken line arrow).

[Optical multilayer film forming process (enlarged view): 340]
Next, in the optical multilayer film forming step 340, the optical multilayer film 2A is laminated and formed. For example, the 2d layer, 2c layer, 2b layer, and 2a layer shown in FIG. 4 are formed in order by a film forming method such as vacuum deposition or sputtering.

[Hydrophobic film forming step: 350]
Next, in the hydrophobic film forming step 350, the hydrophobic film 12 is laminated and formed on the surface of the optical multilayer film 2A formed in the previous step. For example, a water repellent material containing C (carbon) or F (fluorine) or Si (silicon) or a composite water repellent material thereof can be made to a thickness of, for example, 2 nm by a film forming method such as vacuum deposition. Form.

[Hydrophobic membrane removal step: 370]
Finally, in the hydrophobic film removing step 370, the hydrophobic film 12 formed in the inner region (inner center vicinity 21 and intermediate region 24) of the sample loading spot 10 is peeled off. For example, a mask 15 is placed on the upper surface of the sample loading plate 100 by a processing method such as plasma etching, the hydrophobic film 12 is peeled off by a processing method such as plasma etching, and then the mask 15 is removed. The mask 15 is formed with an opening shown as a white portion in FIG. FIG. 5 shows three examples.

In the example shown in FIG. 5A, an opening is formed in the vicinity of the inner center 21 and a concentric mask pattern is formed in the portion with respect to the intermediate region 24 so that the aperture ratio decreases from the center to the outside. ing. The black part acts as a mask that protects from plasma. The portion with respect to the outer peripheral portion 22 is completely masked and the aperture ratio is zero.

In the example shown in FIG. 5B, a radial opening (white portion in the figure) is formed from the portion with respect to the center of the sample loading spot 10 to the outer portion. The portion with respect to the outer peripheral portion 22 is completely masked and the aperture ratio is zero. With such a configuration, it is possible to configure the aperture ratio to decrease from the center to the outside. In this example, the radial opening has a constant width, but the width may become smaller toward the outside. Further, in this example, the central portion where the white openings intersect corresponds to the inner center vicinity 21.

In the example shown in FIG. 5C, an opening is formed in the portion for the inner center vicinity 21 and a mesh-like mask (black portion) is formed in the portion for the intermediate region 24, and the black portion of the mesh is formed. The area ratio is increasing toward the outside. The portion with respect to the outer peripheral portion 22 is completely masked and the aperture ratio is zero. With such a configuration, it is possible to configure the aperture ratio to decrease from the center to the outside.

According to the manufacturing method described above, it is possible to obtain a sample loading spot having high hydrophilicity near the inner center of the spot and gradually becoming hydrophobic toward the outside. In addition, it is possible to provide a method for manufacturing a sample loading plate having a desired reflection color due to an optical multilayer film on the surface of the substrate, and to manufacture a sample loading plate having good sample visibility and a high anchoring effect for a dripping sample. A method can be provided.

Alternatively, in the hydrophobic film forming step 350, the sample loading spot 10 is placed on the sample loading plate 100 with the white portion and the black portion of the mask shown in FIG. 5 reversed, that is, the mask shown in FIG. It is also possible to carry out the hydrophobic film forming step 350 in the covered state, in which case the hydrophobic film removing step 370 becomes unnecessary.

Further, as shown in FIG. 13, it is also possible to expose the substrate 1 by forming a groove 3 on the outer peripheral portion of the intermediate region 24 by a method such as laser marking, and the contrast between the dark blue surface and the white color of the substrate allows the substrate 1 to be exposed. It is also possible to add an effect that the visibility of the sample loading spot 10 is improved and the work of placing the sample is facilitated.

The second embodiment will be described with reference to FIGS. 6 to 10. FIG. 6A is a plan view of the sample loading plate 100 according to the second embodiment. FIG. 6B is a cross-sectional view taken along the cutting line I-I'passing through the center of the sample loading spot 10 shown in FIG. 6A, and is a plan view at the top in order to make the planar positional relationship easy to understand. An enlarged view of is added. In the second embodiment, the substrate 1 is made of a material having a hydrophobic surface (described later). A first metal film 2M is first formed on one side surface of the substrate 1. Next, the optical multilayer film 2A is formed by laminating on the first metal film 2M. The optical multilayer film 2A is made of a dielectric film or a second metal film, and the type and number of layers are not particularly limited, and are formed in the order of, for example, 2d, 2c, 2b, and 2a. The optical multilayer film 2A is hydrophilic and is defined as a first hydrophilic film. The first metal film 2M and the optical multilayer film 2A are formed by a film forming method such as vacuum deposition or sputtering.

As shown in FIG. 6B, in a plane, the vicinity of the inner center 21 is covered with the optical multilayer film 2A and is hydrophilic, and in the intermediate region 24, the optical multilayer film 2A is outside the center of the sample loading spot. The pattern is formed so that the optical multilayer film 2A and the first metal film 2M are peeled off so as to expose the hydrophobic substrate 1 so that the ratio of the area occupied by the optical multilayer film 2A becomes smaller. Is forming. Therefore, the hydrophobicity is gradually increased from the center of the sample loading spot to the outside.

FIG. 7 shows the material and thickness of each layer as the cross-sectional structure of the sample loading plate 100 in Example 2. In the cross-sectional structure shown in FIG. 7, polytetrafluoroethylene (PTFE) is used for the substrate 1. Therefore, the substrate 1 has a hydrophobic surface. The first metal film 2M laminated on the substrate 1 uses Ni as a material and has a film thickness of about 300 nm. Next, the first layer 2d constituting the optical multilayer film 2A uses Al ₂ O _{3 and} has a film thickness of about 80 nm. Ti is used for the second layer 2c, and the film thickness is about 10 nm. The third layer 2b uses SiO _{2 and} has a film thickness of 90 nm. The fourth layer 2a uses Ti and has a film thickness of about 10 nm. With such a film configuration, the surface of the sample loading plate 100 can be dark blue in the visible light wavelength region.

As described above, by preferably combining the first metal film laminated on the substrate 1 and the optical multilayer film 2A, arbitrary reflection characteristics (coloring) utilizing optical interference can be obtained.

Further, by using a white material such as PTFE for the substrate 1, the substrate color exposed as the hydrophobic surface becomes white, the contrast with the optical multilayer film 2A becomes more remarkable, and the visibility of the sample loading spot 10 becomes better. Become. Further, since the crystal of the sample is white, the surface of the sample loading plate can be color-identified from the optical multilayer film 2A, and the presence or absence of loading can be confirmed after crystallization.

[Explanation of Manufacturing Method of Sample Loading Plate 100: FIGS. 9 and 10]
Next, the method of manufacturing the sample loading plate 100 of Example 2 will be described with reference to FIGS. 9 and 10. FIG. 9 is a process diagram showing a method for manufacturing the sample loading plate 100. FIG. 10 is an example of a pattern of the mask 15 used in the manufacturing process.

[Explanation of manufacturing method: Fig. 9]
In FIG. 9, the main steps of 410 to 470 are illustrated and described for the method of manufacturing the sample loading plate 100. Unless otherwise specified in each step, it is natural to perform general operations such as transfer, inspection, cleaning, drying, and annealing necessary for each step, and the description thereof will be omitted.

[Substrate acceptance process: 410]
First, in the substrate receiving step 410, the flatness and surface roughness of the substrate 1 are inspected to confirm that the substrate 1 has a predetermined flatness and surface roughness.

[Substrate surface processing process (enlarged view): 420]
Next, in the substrate surface processing step 420, the substrate 1 is subjected to a lapping process or a polishing process to finish the substrate 1 to a predetermined substrate thickness, surface roughness, and flatness. The main inspection items in this step are the surface roughness and flatness of the substrate. Further, if necessary, a fine structure on the surface may be formed so as to improve the adhesion when forming the metal film in the next step.

[First metal film forming step (enlarged view): 430]
Next, in the first metal film forming step 430, the first metal film 2M is formed. For example, Ni is formed to a thickness of 300 nm by using a film forming method such as vacuum deposition or sputtering. At this time, the irradiation direction of the film-formed particles is preferably vertical in order to make the film as uniform as possible (see the broken line arrow).

[Optical multilayer film forming process (enlarged view): 440]
Next, in the optical multilayer film forming step 440, the optical multilayer film 2A is laminated and formed. For example, the 2d layer, 2c layer, 2b layer, and 2a layer shown in FIG. 4 are formed in order by a film forming method such as vacuum deposition or sputtering.

[Removal step of optical multilayer film 2A and first metal film 2M: 470]
Finally, in the removal step 470 of the optical multilayer film 2A and the first metal film 2M, the optical multilayer film 2A and the first metal formed in the inner region (inner center vicinity 21 and intermediate region 24) of the sample loading spot 10 are formed. The film 2M is peeled off. For example, a mask 15 is placed on the upper surface of the sample loading plate 100 by a processing method such as plasma etching, and the optical multilayer film 2A and the first metal film 2M are peeled off by a processing method such as plasma etching. The step is to remove the mask 15. The mask 15 is formed with an opening shown as a white portion in FIG. FIG. 10 shows three examples.

In the example shown in FIG. 10A, a black mask portion is formed near the inner center 21 and a concentric mask pattern is formed in the portion with respect to the intermediate region 24 so that the aperture ratio increases from the center to the outside. Has been done. The black part acts as a mask that protects from plasma. The portion with respect to the outer peripheral portion 22 is an opening except for a portion having a thin crosspiece for connecting masks at adjacent sample loading spots to maintain a positional relationship.

In the example shown in FIG. 10B, a radial mask portion (black portion in the figure) is formed from a portion with respect to the center of the sample loading spot 10 toward an outer portion. The portion with respect to the outer peripheral portion 22 is an opening except for a portion having a thin crosspiece as in FIG. 5A. With such a configuration, it is possible to configure the aperture ratio to increase from the center to the outside. In this example, the radial mask portion has a constant width, but the width may become smaller toward the outside. Further, in this example, the central portion where the white openings intersect corresponds to the inner center vicinity 21.

In the example shown in FIG. 10C, the portion with respect to the inner center vicinity 21 is a black mask and the aperture ratio is zero, and the portion with respect to the intermediate region 24 is formed with a mesh-like opening (white portion). The area ratio of the mesh opening (white part) is increasing toward the outside. The portion with respect to the outer peripheral portion 22 is an opening except for a portion having a thin crosspiece. With such a configuration, the aperture ratio can be increased from the center to the outside.

According to the manufacturing method described above, it is possible to obtain a sample loading spot having high hydrophilicity near the inner center of the spot and gradually becoming hydrophobic toward the outside. In addition, it is possible to provide a method for manufacturing a sample loading plate having a desired reflection color due to an optical multilayer film on the surface of the substrate, and to manufacture a sample loading plate having good sample visibility and a high anchoring effect for a dripping sample. A method can be provided.

Alternatively, in the first metal film forming step 430 and the optical multilayer film forming step 440, the sample loading spot 10 is reversed from the white portion and the black portion of the mask shown in FIG. 10 above the sample loading plate 100. It is also possible to carry out the first metal film forming step 430 and the optical multilayer film forming step 440 with the mask, that is, the mask shown in FIG. 5 covered, in which case the optical multilayer film 2A and the first The removal step 470 of the metal film 2M of 1 becomes unnecessary.

Although various embodiments of the sample loading plate and the manufacturing method thereof have been described in detail above, the present invention is not limited to such embodiments and manufacturing methods, and the present invention is not limited to such embodiments and manufacturing methods. It can be arbitrarily changed, added or deleted without departing from the idea of the invention.

[Explanation of coloring and visibility of sample loading plate: FIG. 11]
Next, the coloring of the sample loading plate will be described with reference to FIG. FIG. 11 is a schematic diagram illustrating light interference when an optical multilayer film is formed on a substrate.

In FIG. 11, for the sake of explanation, for example, dielectric films 2a, 2b, 2c, and 2d are laminated and formed as an optical multilayer film. Arbitrary reflection characteristics (coloring) can be obtained by adjusting the material (refractive index), thickness, and number of layers of each layer, but here, only a schematic diagram is used to explain the principle. (Generally, by alternately laminating a dielectric film having a high refractive index and a dielectric film having a low refractive index as a pair with a thickness of 1/4 wavelength, the reflected wave from the interface of each layer is phased by the interference action of light. It is said that a highly efficient reflection function can be obtained by additionally overlapping.)

The incident light P incident on the optical multilayer film from the air layer 90 first generates a reflected wave 2aR at the interface between the air and the dielectric film 2a. Similarly, the reflected waves 2bR, 2cR, 2dR, and 1R are generated at the interface of each layer. The reflections from each interface are added together to form a reflected wave R. The reflected wave R can obtain arbitrary reflection characteristics (coloring) by changing the material (refractive index), film thickness, and number of film layers of each layer. Various reflection characteristics can be obtained by mixing a metal film with the dielectric film.

Based on this principle, as a result of selecting specific film quality and film thickness as shown in FIG. 2 in Example 1 and FIG. 7 in Example 2, the reflection characteristic of the sample loading plate 100 is in the wavelength region of visible light. At W (about 380 nm to about 780 nm), the reflectance is low as a whole, but it has a peak on the side where the wavelength is small, that is, a large amount of dark blue light is reflected, and the surface of the plate appears to be colored dark blue.

Further, an arbitrary color can be created by the first metal film 2M and the optical multilayer film 2A laminated on the substrate. As a result, the visibility of the loaded sample can be improved, and the efficiency of the sample dropping operation is improved. In addition, by making sample loading plates of various colors and color-coding them, it becomes easy to store and manage the samples.

[Explanation of analysis operation by mass spectrometer: FIG. 3, FIG. 8, FIG. 12]
Next, the operation of performing mass spectrometry of the sample will be described with reference to FIGS. 3, 8, and 12. Here, the sample loading plate and the part related to the ionization of the sample will be mainly described, and the other parts will be described in principle and the details will be omitted. 3 and 8 are schematic views showing a state in which the sample 200 is loaded on the sample loading plate 100 described above, FIG. 3 shows a state in Example 1, and FIG. 8 shows a state in Example 2. FIG. 12 is a schematic view showing a state in which a sample loading plate on which the sample 200 is loaded is placed on the mass spectrometer 300.

In both Example 1 (FIG. 3) and Example 2 (FIG. 8), a sample 200 obtained by mixing an object to be analyzed and a matrix and liquefied with a solvent is dropped onto a sample loading spot to evaporate the solvent, and a cross section of the dried state is obtained. It is shown in the figure. A predetermined amount of the sample 200 is dropped onto the sample loading spot 10 near the inner center 21 (see FIGS. 1 and 6) by an instrument (not shown). The dropped sample 200 tries to spread radially due to gravity and surface tension, but is attracted to the hydrophilic surface and traps the sample in the area obtained by adding the inner center vicinity 21 and the intermediate region 24. Since the intermediate region 24 is around and has a hydrophilic surface as compared with the case where only the inner center vicinity 21 is a hydrophilic surface, the droplets of the sample can be reliably trapped in a wider area.

Further, when the hydrophilic surface is formed by the optical multilayer film, the reflectance of light can be adjusted. In this embodiment, the color of the substrate is dark blue, and the color of the substrate is alumina or white in PTFE. Therefore, the exposed portion of the sample loading spot 10 on the substrate serves as a mark when the sample is loaded. Since the crystal of the sample is white, the color can be distinguished from the surface of the sample loading plate, and the presence or absence of loading can be confirmed after crystallization.

After the loading of the sample 200 to be analyzed is completed, each sample 200 is dried in that state. As the volume of the sample gradually decreases, the hydrophilic surface is attracted to the vicinity of the inner center where it is most exposed, and it becomes possible to retain more dried and crystallized sample near the inner center. It is a structure that gradually changes from a hydrophilic surface to a hydrophobic surface from the center of the sample loading spot that gives such a result toward the outside. In manufacturing a sample loading plate having this structure, it is possible to easily manufacture the sample loading plate by removing the outermost film of the sample loading plate using one mask.

Next, FIG. 12 shows a schematic view of the mass spectrometer 300, and the sample loading plate 100 on which the sample 200 is loaded is placed on the mass spectrometer 300 and fixed by a fixing portion (not shown). Actually, the sample 200 loaded on a plurality of spots has a mechanism that allows each sample to move in the X and Y directions and stop at a predetermined position. However, for the sake of simplicity, one sample loading spot is used here. Will be described.

In the mass spectrometer 300 shown in FIG. 12, the sample loading plate 100 is placed on the left side and is detachably fixed by a clamp portion (not shown). Further, a voltage application portion (not shown) can conduct electricity to the sample loading plate 100. Further, a laser light source 220 that irradiates the sample 200 with a laser beam 220a, and an ion accelerator 230 that accelerates an object to be analyzed (200a, 200b, 200c,) separated from the sample 200 and ionized by the irradiation of the laser beam. It is provided with an ion trap unit 231 for trapping ions, a mass separation unit 232 for forming an ion flight space and performing mass separation of each ion, and an ion detection unit 240 for detecting each ion reached by mass separation in time series. ing.

Here, it is assumed that the polarity of the ions of the object to be analyzed is positive (positive charge). When the mass spectrometry starts, the laser light source 220 irradiates the sample 200 to be measured with the laser beam 220a for a predetermined time. At the same time, a positive voltage V1 is applied to the first metal film 2M of the sample loading plate 100 and the metal film (2a, 2c) in the optical multilayer film from a voltage application portion (not shown), and a positive voltage is applied to the sample 200 with respect to the sample 200. Is effectively given to. At the same time, a negative voltage V2 is applied to the first grid of the ion trap unit 231.

At this time, the matrix contained in the sample 200 is vaporized together with the object to be analyzed, and the object to be analyzed is desorbed and ionized. Then, a positive voltage V1 is applied to the object to be analyzed, and a downward-grading electric field is generated toward the ion trap unit 231 to which the negative voltage V2 is applied. Therefore, the desorbed and ionized object to be analyzed is an ion acceleration unit. At 230, it is accelerated toward the ion trap unit 231. The object to be desorbed and ionized in this way is sent from the ion trap unit 231 to the mass separation unit (flight space) 232, separated by the difference in mass during flight, and has a time difference of 200c, 200b, 200a. It reaches the ion detection unit in order. Then, the data detected by the ion detection unit 240 is analyzed by an analyzer (not shown) to perform mass spectrometry on the object to be analyzed. As a result, sample identification is performed at high speed and with high accuracy.

The embodiments and effects of the present invention have been described above based on Examples 1 and 2. In Example 1, an example in which the ceramic Al ₂ O ₃ is used for the substrate 1 has been described, but the present invention is not limited to this, and other ceramic materials, porcelain and ceramic composite materials, glass, Si, plastic, and the like may be used. Good. Further, an example in which Ni, Ti, and Al are used as the metal film of the first metal film 2M and the optical multilayer film 2A has been described, but the present invention is not limited to this, and other metals such as chromium and gold may be used. Further, an example in which Al ₂ O ₃ , SiO ₂ , and TiO ₂ are used as the material of the dielectric film has been described, but the present invention is not limited to this, and other dielectric materials such as MgO, MgF ₂ , and ZrO ₂ may be used.

In the second embodiment, the substrate 1 is made of a resin PTFE to have a hydrophobic surface, but the present invention is not limited to this, and the substrate is made of, for example, a material such as ceramics or metal, and the hydrophobic surface is formed on the surface thereof. May be used.

Further, in this embodiment, the metal film and the optical multilayer film are formed only on one side surface of the substrate, but it is more convenient to form the metal film or the optical multilayer film on both side surfaces of the substrate depending on the film forming method. In some cases, a metal film and an optical multilayer film may be formed on both side surfaces of the substrate, or either a metal film or an optical multilayer film may be formed on the surface on the side where the sample is not loaded, or even in a plane. It may be partially formed.

1 Substrate 2A Optical multilayer film (first hydrophilic film)
2M 1st metal film 2a, 2b, 2c, 2d Dielectric film or 2nd metal film 10 Sample loading spot 12 Hydrophobic film 15 Mask 20 Marginal part of sample loading plate 21 Near the inner center (of sample loading spot) 22 Outer peripheral part (of sample loading spot) 24 Intermediate area (of sample loading spot) 30 columns Address mark 40 rows Address mark 50 Serial number 60 Barcode 100 Sample loading plate 200 Sample 200a, 200b, 200c Ionized sample 220 Laser light source 220a Laser light 230 Ion acceleration part 231 Ion trap part 232 Mass separation part (flying space)
240 Ion detector 300 MALDI mass spectrometer

Claims (10)

A sample loading plate used for mass spectrometry of the MALDI method and having at least one sample loading spot for loading a sample on a substrate.
A hydrophobic surface is formed on the edge of the sample loading plate, which is an outer region of the sample loading spot.
The sample loading spot includes a vicinity of the inner center and an intermediate region from the vicinity of the inner center to the edge.
A hydrophilic surface is formed near the inner center of the sample loading spot.
A hydrophilic surface and a hydrophobic surface are partially formed in the intermediate region of the sample loading spot, and the area ratio of the hydrophobic surface increases from the vicinity of the inner center toward the edge portion. A sample loading plate, characterized in that it is formed . The sample loading plate according to claim 1, wherein the substrate is a resin, and the hydrophobic surface formed in the peripheral portion and the intermediate region is exposed. The sample loading plate according to claim 1 or 2, wherein the hydrophilic surface formed in the vicinity of the inner center and the intermediate region exposes the optical multilayer film formed on the substrate. The sample loading plate according to claim 1, wherein the substrate is ceramics or metal, and the hydrophilic surfaces formed in the vicinity of the inner center and the intermediate region are exposed. A method for manufacturing a sample loading plate used for mass spectrometry of the MALDI method and having at least one sample loading spot for loading a sample on a substrate.
The process of making the substrate surface a hydrophilic surface,
A step of forming a hydrophobic film on the hydrophilic surface to form a hydrophobic surface,
An opening is formed in a portion of the sample loading spot in the vicinity of the inner center from the edge portion, and no opening is formed in the portion of the sample loading spot in the vicinity of the inner center. A mask having an opening formed so that the proportion of the opening decreases toward the outside from the center of the sample loading spot is placed on the surface of the sample loading plate at the portion for the intermediate region between the two. And the process to do
A step of removing the hydrophobic film on the surface of the sample loading spot using the mask, and
A method for manufacturing a sample loading plate, which comprises a step of removing the mask. The mask has a plurality of concentric openings with respect to the center of the sample loading spot, is open at a portion near the inner center, and has no opening at a portion with respect to the edge portion. The method for manufacturing a sample loading plate according to claim 5, wherein the ratio of the opening area to the intermediate region gradually decreases from the center of the sample loading spot toward the outside. 5. The mask is characterized by having a plurality of linear openings extending radially from the center of the sample loading spot and having a constant width or a width that decreases outward from the center. The method for manufacturing a sample loading plate according to. A method for manufacturing a sample loading plate used for mass spectrometry of the MALDI method and having at least one sample loading spot for loading a sample on a substrate.
The process of making the substrate surface a hydrophobic surface,
A step of forming a hydrophilic film on the hydrophobic surface to form a hydrophilic surface,
An opening is not formed in a portion of the sample loading spot in the vicinity of the inner center from the edge portion, and an opening is formed in the portion of the sample loading spot in the vicinity of the inner center, and the vicinity of the inner center and the margin portion are formed. A mask having an opening formed so that the proportion of the opening increases toward the outside from the center of the sample loading spot is placed on the surface of the sample loading plate at the portion between and the intermediate region. The process of placing and
A step of removing the hydrophilic film on the surface of the sample loading spot using the mask, and
A method for manufacturing a sample loading plate, which comprises a step of removing the mask. The mask has a plurality of concentric openings with respect to the center of the sample loading spot, has no opening in the portion near the inner center, and has an opening in the portion with respect to the edge portion. The method for manufacturing a sample loading plate according to claim 8, wherein the ratio of the opening area to the intermediate region gradually increases from the center of the sample loading spot toward the outside. The mask has non-openings at sites for a plurality of linear regions extending radially from the center of the sample loading spot and having a constant width or a width that decreases outward from the center. The method for manufacturing a sample loading plate according to claim 8.

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