JP2009236489A - Sample target used in mass analyzing method, its manufacturing method and mass analyzer using sample target - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a sample target which enhances the measuring sensitivity of a low-molecular compound and is able to ionize a substance with a high-molecular weight of above 30,000 in mass analysis enabling the ionization of a sample without using a matrix, its manufacturing method and a mass analyzer using the sample target. <P>SOLUTION: The sample target is used for holding the sample when the sample is ionized by the irradiation with a laser beam to perform mass analysis and has a surface, which receives the irradiation with the laser beam and has a plurality of recessed parts repeatedly formed thereto, as a sample holding surface. The surface of the sample holding surface excepting a part of the inner surfaces of the recessed parts is covered with a metal and/or a semiconductor and a part uncovered with the metal and/or the semiconductor is present on the inner surfaces of the recessed parts. <P>COPYRIGHT: (C)2010,JPO&INPIT

Description

  The present invention relates to a sample target used for mass spectrometry, a method for producing the same, and a mass spectrometer using the sample target, and particularly enables ionization of a high molecular weight sample even when a matrix is not used. The present invention relates to a sample target to be manufactured, a manufacturing method thereof, and a mass spectrometer using the sample target.

  The mass spectrometry is an analysis method in which a sample is ionized, the ratio of the mass of the sample or fragment ions of the sample and the charge (hereinafter referred to as m / z value) is measured, and the molecular weight of the sample is examined. Among them, the matrix-assisted laser desorption / ionization mass spectrometry (MALDI) method mixes a sample with a low molecular weight organic compound called a matrix and irradiates the sample with a laser. This is a method of ionizing the. In this method, the energy of the laser absorbed by the matrix is transmitted to the sample, so that the sample can be ionized well.

  The MALDI method can ionize a thermally unstable substance or a high molecular weight substance, and can ionize a sample “softly” as compared with other ionization techniques. Therefore, this method is widely used for mass spectrometry of various substances such as biopolymers, endocrine disruptors, synthetic polymers, and metal complexes.

  However, since the MALDI method uses a matrix of an organic compound, it may be difficult to analyze sample ions due to related ions derived from the matrix. Specifically, when an organic compound matrix is used, matrix-related ions such as ions of the matrix molecules, ions of clusters in which the matrix molecules are bonded by hydrogen bonds, and fragment ions generated by decomposition of the matrix molecules are observed. Therefore, it is often difficult to analyze sample ions.

  Therefore, conventionally, various techniques for avoiding the interference of the matrix-related ions have been proposed. Specifically, as a technique for immobilizing matrix molecules so as not to generate matrix-related ions, for example, a technique for immobilizing a matrix such as α-cyano-4-hydroxycinnamic acid or cinnamamide on Sepharose beads, a target A technique of forming a self-assembled monolayer of methyl-N- (4-mercaptophenyl-carbamate) as a matrix on the surface of a certain gold, by a sol-gel method, 2,5-dihydroxybenzoic acid (DHB) as a matrix A technique for fixing the resin in a silicon polymer sheet is known. However, the method of immobilizing matrix molecules as described above has a problem that the detection sensitivity and durability are not sufficient in practice. Also, there is a problem that noise due to fragment ions cannot be avoided during detection.

  Therefore, recently, a technique that does not use a matrix has been proposed. Specifically, a technique using a semiconductor substrate having a multi-hole surface (described as a porous light-absorbing semiconductor substrate in the literature) as a sample target is disclosed (for example, see Patent Document 1). In this sample target, the sample holding surface of the semiconductor substrate is processed to have a porous structure, that is, a fine uneven structure. This document reports that when a sample is applied to such a sample holding surface and the sample is irradiated with laser light, a high molecular weight substance is ionized without a matrix. This method is named DIOS (Desorption / Ionization on Porous Silicon) method.

  In addition, as a technique that enables ionization even when a matrix is not used, the present inventors have developed a surface having a fine and regular concavo-convex structure on the order of nanometers to several tens of micrometers produced by lithography. And a sample target having a sample holding surface in which a surface having a fine concavo-convex structure on the order of nanometers to several tens of micrometers is coated with a metal. In comparison, it has been found that ionization efficiency can be improved and more stable ionization is possible (see, for example, Patent Document 2). Note that a method of ionizing a sample without using a matrix using a sample target having a surface having a concavo-convex structure on the order of nanometers to several tens of micrometers as a sample holding surface is described in “Surface-Assisted Laser Desorption / Ionization Mass”. Analysis method (SALDI) ".

  Furthermore, the present inventors have examined a sample target having a surface made of various materials having a concavo-convex structure on the order of submicrometers as a sample holding surface. As one of such materials, porous alumina is used. It has also been found that ionization is possible without using a matrix even when a sample holding surface coated with gold or platinum is used (see, for example, Non-Patent Document 1).

Further, the inventors of the present invention have provided a sample target having a sample holding surface having a large number of pores opened on the surface that is irradiated with the laser beam, and the pore diameter of the pores is 30 nm or more and less than 5 μm and is fine. By using a sample target having a pore depth / (pore cycle−pore diameter) of 2 or more and 50 or less and the surface of the sample holding surface is coated with a metal and / or a semiconductor, a molecular weight can be obtained without using a matrix. Has been found to be capable of ionizing samples exceeding 10,000 (for example, see Patent Document 3).
US Pat. No. 6,288,390 (published September 11, 2001) International Publication No. 2005/083418 Pamphlet (published September 9, 2005) International Publication No. 2007/046162 pamphlet (released on April 26, 2007) Shoji Okuno, Ryuichi Arakawa, Kazumasa Okamoto, Yoshinori Matsui, Shu Seki, Takahiro Kozawa, Seiichi Tagawa, Yoshinao Wada, Matrix-free laser desorption / ionization of peptides on sub-micrometer structures: Grooves on silicon and metal-coated porous alumina, 53rd ASMS Conference on Mass Spectrometry and Allied Topics, (San Antonio, Texas, USA), Proceedings (Published on April 15, 2005 on the web)

  However, the conventional laser desorption ionization mass spectrometry without using a matrix cannot ionize a high molecular weight substance having a molecular weight exceeding 30000. Further, further improvement in measurement sensitivity is desired even for low molecular weight compounds.

  That is, the conventional laser desorption ionization mass spectrometry by DIOS method disclosed in Patent Document 1 is effective for ionizing a substance having a molecular weight of 3000 or less, but cannot ionize a substance having a molecular weight exceeding 10,000. .

  In addition, a sample target provided with a sample holding surface on a surface having a regular concavo-convex structure manufactured by a lithography method disclosed in Patent Document 2 or a sample holding surface in which a surface having a fine concavo-convex structure is coated with a metal Even if a laser is used without using a matrix, a large substance having a molecular weight exceeding 10,000 cannot be ionized.

  Using a sample target disclosed in Patent Document 3 having a pore diameter smaller than a certain size and having a pore depth in a predetermined range / (pore cycle-pore diameter), laser irradiation is performed without using a matrix. Even in this case, a substance having a molecular weight exceeding 10,000 can be ionized, but a substance having a high molecular weight having a molecular weight exceeding 30,000 cannot be ionized.

  The present invention has been made in view of the above problems, and its object is to enable ionization of a high molecular weight substance having a molecular weight exceeding 30000 in mass spectrometry that enables ionization of a sample without using a matrix. At the same time, it is to provide a sample target and a method for producing the sample target in which the measurement sensitivity of the low molecular compound is further improved, and a mass spectrometer using the sample target.

  In order to solve the above problems, the sample target according to the present invention is used for holding a sample when ionizing the sample by laser beam irradiation to perform mass spectrometry, and is applied to the surface receiving the laser beam irradiation. A sample target having a surface on which a plurality of recesses to be opened are repeatedly formed as a sample holding surface, and the interval between adjacent recesses is 1 nm or more and less than 30 μm, and a part of the inner surface of the recess is formed. The surface of the sample holding surface to be removed is covered with metal and / or semiconductor, and a portion not covered with metal and / or semiconductor is present on the inner surface of the recess.

  Thereby, ionization of a substance having a molecular weight exceeding 30000 becomes possible. Moreover, the measurement of the low molecular weight compound which has been the main analysis target of the conventional SALDI method can be performed with higher sensitivity.

  In the sample target according to the present invention, the portion of the inner surface of the recess that is covered with metal and / or semiconductor is the upper portion of the side wall including the upper end of the side wall of the recess, It is preferable that the distance between the lower end of the portion being formed is 0 or more on average and the width of the recess × 1.73 or less.

  In the sample target according to the present invention, it is preferable that the plurality of concave portions are regularly and repeatedly formed.

  By forming each of the concave portions regularly and repeatedly, there is little variation in the structure having the concave portions, so that the ionization performance can be further stabilized.

  In the sample target according to the present invention, the recess is preferably a groove or a pore.

  In the sample target according to the present invention, the sample holding surface is preferably made of porous alumina.

  Porous alumina has a plurality of regularly formed recesses, and can obtain a structure having a preferable pore depth / (pore cycle−pore diameter) for the sample target. Furthermore, porous alumina can be suitably used in the present invention because the pore diameter, pore depth, and pore cycle can be controlled by changing the anodizing conditions.

  In the sample target according to the present invention, the sample holding surface has a negative-type first structure in which the concavo-convex structure of the first structure is transferred using the first structure having the concavo-convex structure in which fine concave portions are repeatedly formed as a mold. A sample holding surface having a concavo-convex structure having the same shape as the concavo-convex structure of the first structure, in which two structures are produced and the concavo-convex structure is transferred using the negative second structure as a mold. Preferably there is. The first structure is preferably porous alumina.

  Thereby, the sample holding surface which has the same structure as the 1st structure which has an uneven structure suitable for a sample holding surface can be manufactured with a desired material.

  In the sample target according to the present invention, the sample holding surface preferably contains a resin or a ceramic.

  In the sample target according to the present invention, the metal is preferably platinum (Pt) or gold (Au).

In the sample target of the present invention, the _{semiconductor, Si, Ge, SiC, GaP} , GaAs, InP, Si 1-X Ge X (0 <X <1), SnO 2, ZnO, In 2 O 3, or, A mixture of two or more of these or carbon is preferable.

  In order to solve the above problems, the sample target manufacturing method according to the present invention is used for holding a sample when ionizing the sample by laser beam irradiation and performing mass spectrometry, and irradiating the laser beam. A method of manufacturing a sample target comprising, as a sample holding surface, a surface in which a plurality of recesses that are open on the receiving surface are repeatedly formed, wherein the surface of the sample holding surface is made of metal and / or semiconductor on the inner surface of the recess It is characterized by including a coating step of coating with metal and / or semiconductor so that there is an uncoated portion.

  Thereby, ionization of a substance having a molecular weight exceeding 30000 becomes possible. Moreover, the measurement of the low molecular weight compound which has been the main analysis target of the conventional SALDI method can be performed with higher sensitivity.

  In the sample target manufacturing method according to the present invention, the coating step uses physical vapor deposition and is inclined obliquely from the center line of the recess extending in the thickness direction from the surface of the sample holding surface or from the direction of the center surface. From the angle, it is preferable to deposit the metal and / or semiconductor.

  Thereby, ionization of a substance having a molecular weight exceeding 30000 becomes possible. Moreover, the measurement of the low molecular weight compound which has been the main analysis target of the conventional SALDI method can be performed with higher sensitivity.

  In the sample target manufacturing method according to the present invention, the physical vapor deposition is preferably sputtering, thermal vapor deposition, electron beam vapor deposition, or vacuum vapor deposition.

  In the sample target manufacturing method according to the present invention, after the coating step, a portion of the inner surface of the recess that is not covered with metal and / or semiconductor is etched to increase the diameter of the recess of the portion. It is preferable to include a process.

  Thereby, ionization performance can be further improved.

  In the sample target manufacturing method according to the present invention, porous alumina can be suitably used as the sample holding surface.

  The sample target manufacturing method according to the present invention uses a first structure having a concavo-convex structure in which fine concave portions are repeatedly formed as a mold before the step of coating the surface of the sample holding surface with a metal and / or a semiconductor. Using the second structure of the negative structure obtained by transferring the concavo-convex structure of the first structure using the negative second structure obtained in the process as a mold to transfer the concavo-convex structure. And obtaining a sample holding surface having a concavo-convex structure having the same shape as the concavo-convex structure of the first structure on the surface. The first structure is preferably porous alumina.

  In the method of manufacturing a sample target according to the present invention, before the step of coating the surface of the sample holding surface with metal and / or semiconductor, the space between the recesses is 1 nm or more and 30 μm on the surface of the substrate using a lithography technique. The step of forming the sample holding surface on the surface may be included by repeatedly forming the recess so as to be less.

  A mass spectrometer according to the present invention uses the sample target. The mass spectrometer is preferably a laser desorption ionization mass spectrometer that irradiates a sample to be measured with laser light to ionize the sample and measure its molecular weight.

  As described above, the sample target according to the present invention is a sample target having a surface on which a plurality of fine recesses that are opened on the surface that is irradiated with laser light are repeatedly formed as a sample holding surface, The surface of the sample holding surface excluding a part of the inner surface of the recess is covered with a metal and / or a semiconductor, and a portion not covered with the metal and / or semiconductor exists on the inner surface of the recess. Therefore, even when ionization is performed without using a matrix when performing mass spectrometry, high ionization performance can be realized. Thereby, ionization of a substance having a molecular weight exceeding 30000 becomes possible. Moreover, the measurement of the low molecular weight compound which has been the main analysis target of the conventional SALDI method can be performed with higher sensitivity.

  In addition, as described above, the method of manufacturing the sample target according to the present invention includes the metal and the surface of the sample holding surface so that the inner surface of the concave portion is not covered with metal and / or semiconductor. Since the structure including the coating step of coating with a semiconductor is provided, high ionization performance can be realized even when ionization is performed without using a matrix when performing mass spectrometry. Thereby, ionization of a substance having a molecular weight exceeding 30000 becomes possible. Moreover, the measurement of the low molecular weight compound which has been the main analysis target of the conventional SALDI method can be performed with higher sensitivity.

  The present invention will be described in detail below, but the present invention is not limited to the following description.

  As a result of intensive studies to solve the above problems, the inventors of the present invention coat a metal and / or semiconductor on the surface of a sample holding surface in which a plurality of minute recesses are repeatedly formed in a direction perpendicular to the sample holding surface. In some cases, the metal and / or semiconductor is deposited at an angle perpendicular to the sample holding surface as has been conventionally done, and the metal and the metal target are inclined at an angle with respect to the sample holding surface. In the sample target obtained by vapor-depositing a semiconductor, it has been found that the ionization performance of a low molecular compound is remarkably improved. Therefore, when a high molecular weight material having a molecular weight exceeding 30000 is ionized, a sample target obtained by depositing metal and / or semiconductor at an angle perpendicular to the sample holding surface cannot ionize such material. It has been found for the first time that a large substance having a molecular weight exceeding 30000 can be ionized in a sample target obtained by vapor-depositing a metal and / or semiconductor at an angle with respect to the surface. In the conventional vapor deposition at an angle perpendicular to the sample holding surface, metal and / or semiconductor penetrates into the inside of the recess, and as a result, the inner surface of the recess is covered, but obliquely with respect to the sample holding surface. In the sample target coated with metal and / or semiconductor deposited at such an angle, only the upper part of the inner surface of the recess is covered with metal and / or semiconductor, so that the inner surface of the recess is covered with metal and / or semiconductor. It has been found that the presence of a non-existing part is related to the ionization performance, and the present invention has been completed.

  That is, the sample target according to the present invention is used for holding a sample when ionizing the sample by laser light irradiation and performing mass spectrometry, and has a plurality of recesses opened on the surface receiving the laser light irradiation. A sample target having a repeatedly formed surface as a sample holding surface, and the interval between adjacent recesses is 1 nm or more and less than 30 μm, and the sample holding surface excluding a part of the inner surface of the recess The surface is covered with metal and / or semiconductor, and there is a portion not covered with metal and / or semiconductor on the inner surface of the recess.

  The reason why the ionization performance is remarkably improved by the presence of a portion not coated with metal and / or semiconductor on the inner surface of the concave portion of the sample holding surface is that the high molecular weight substance exceeding 30000 can be ionized. Although it is not clear, one possibility is that water molecules adsorbed on the inner surface of the concave portion of the sample holding surface that is not coated with metal and / or semiconductor evaporate by laser irradiation. It can be considered that they are involved in improvement. That is, in a part not covered with metal and / or semiconductor, water molecules adsorbed on the part are evaporated by laser irradiation to improve ionization performance, which is considered as one of the reasons for improving ionization performance. .

  Hereinafter, with respect to the sample target and the manufacturing method thereof according to the present invention and the mass spectrometer using the sample target, (I) the sample target, (II) the method of manufacturing the sample target, and (III) the use of the present invention (mass analysis) Device) will be described in this order.

(I) Sample target The sample target according to the present invention is used in a laser desorption / ionization mass spectrometer that performs mass analysis by ionizing a sample by irradiation with laser light, and serves as a sample stage on which a sample to be analyzed is placed. It fulfills its function.

  Such a sample target may be provided with a sample holding surface that is a surface for holding a sample, and the configuration, shape, material, and the like of the portion other than the sample holding surface are not particularly limited.

  The sample holding surface of the sample target according to the present invention is a surface for holding a sample to be analyzed, and is irradiated with laser light while holding the sample.

  The sample target according to the present invention includes a surface on which a plurality of recesses opened on the surface that is irradiated with laser light are repeatedly formed as a sample holding surface, and the interval between adjacent recesses is 1 nm or more and less than 30 μm. The surface of the sample holding surface excluding a part of the inner surface of the recess is coated with metal and / or semiconductor, and the inner surface of the recess is not covered with metal and / or semiconductor. Exists.

(I-1) Shape of Sample Holding Surface The sample holding surface of the sample target according to the present invention is a surface in which a plurality of recesses that are open on the surface that is irradiated with laser light is repeatedly formed, and the interval between adjacent recesses Is 1 nm or more and less than 30 μm.

  Here, the interval between adjacent recesses refers to the distance between the center of the width of the groove and the center of the width of the groove adjacent to the recess when the recess is, for example, a groove. In some cases, it refers to the distance between the center line of the pore and the center line of the pore adjacent thereto.

  The surface of the sample holding surface excluding a part of the inner surface of the recess is used as a sample holding surface using a surface where a plurality of recesses are repeatedly formed such that the interval between adjacent recesses is 1 nm or more and less than 30 μm. The measurement sensitivity of the low molecular weight compound can be remarkably improved by coating with metal and / or semiconductor, and the presence of a portion not coated with metal and / or semiconductor on the inner surface of the recess, and the molecular weight is 30000. It becomes possible to ionize a substance having a high molecular weight exceeding. Since the interval between the recesses is narrower than about 30 μm, ionization of the measurement sample in mass spectrometry can be performed satisfactorily. Moreover, it can avoid that the intensity | strength of a sample target falls because the space | interval of each recessed part is 1 nm or more.

  Furthermore, in order to further improve the function as a sample target for mass spectrometry, the interval between the adjacent recesses is more preferably 1 nm or more and less than 10 μm, and preferably 10 nm or more and less than 10 μm. More preferably, it is 10 nm or more and less than 500 nm, particularly preferably 10 nm or more and less than 300 nm. Thereby, ionization of the measurement sample in mass spectrometry can be performed more satisfactorily.

  Further, the interval between the recesses may be regular or irregular. However, in order to further improve the function as a sample target for mass spectrometry, regularity is more preferable. When the interval between the recesses is regular, the ionization performance is more stable because there is little variation in the structure having the recesses.

  The depth of the recess may be about 1 nm or more and less than 30 μm. However, in order to further improve the function as a sample target for mass spectrometry, it is more preferably 1 nm or more and less than 5 μm, further preferably 10 nm or more and less than 1 μm, and particularly preferably 50 nm or more and less than 500 nm. Preferably, it is 100 nm or more and less than 500 nm. Further, the depth of the concave portion may vary or may be uniform. However, in order to further improve the function as a sample target for mass spectrometry, the depth of the concave portion is preferably uniform. When the depth of the recess is uniform, the ionization performance is more stable because there is little variation in the structure having the recess.

  In addition, when the space | interval of the said recessed part and the depth of the said recessed part are not uniform, it is set as these values using an average value.

  The specific shape of the recess is not particularly limited, and may be any shape. Moreover, the shape of a recessed part is not fixed, and the recessed part of various shapes may be mixed. However, in order to further improve the function as a sample target for mass spectrometry, it is preferable that the shape of the recess is constant. Examples of such shapes include shapes such as grooves and pores. Further, the shape of the groove and the pore is not particularly limited, and may be any shape. For example, a straight groove; a curved groove; an arc-shaped groove; Examples include pores; pores having an elliptical cross section; polygonal pores such as a triangle, quadrangle, and pentagon.

  Further, the concave portion may be formed perpendicular to the sample holding surface or may be formed to have an inclination. That the recess is formed perpendicular to the sample holding surface means that the angle formed by the recess and the sample holding surface, that is, the angle formed by the sample holding surface and the center line or the center plane of the recess is vertical. Say.

  Further, the concavo-convex structure may be formed on the entire sample holding surface or may be partially formed on the sample holding surface.

  As described above, it is more preferable that the surface of the sample holding surface of the sample target of the present invention has a structure in which a plurality of concave portions are regularly and repeatedly formed.

<Concavity having groove shape>
FIG. 3 shows a schematic diagram of the groove shape of the sample target when the concave portion is a groove. 3, (a) is a perspective view showing a part of the sample target, (b) is a plan view seen from above the sample holding surface (in the direction of arrow A in (a)), and (c) is a groove shape. It is sectional drawing (sectional drawing seen from the arrow B direction in (a)). Here, the interval between the recesses (grooves) means the size of the portion shown by C in FIG. 3, and the width of the recesses (grooves) means the size of the portion shown by D in FIG. The depth of the recess (groove) means the size of the portion indicated by E in FIG. In FIG. 3, the alternate long and short dash line is the center line of the recess (groove). Further, when a surface formed from the center line exists as in the case of the groove, this surface is set as the center surface of the recess. Moreover, when the space | interval of a recessed part (groove), the width | variety of the said recessed part (groove), and the depth of a recessed part (groove) are not uniform, it is set as these values using average value.

  In the groove-type sample target, if the interval between the grooves is less than 30 μm, more preferably less than 1 μm, the sample placed on the sample target can be ionized well when mass spectrometry is performed. Further, if the groove interval is 1 nm or more, more preferably 10 nm or more, it is possible to perform processing without using advanced techniques in the current microfabrication technique. In order to better ionize the measurement sample, it is more preferable that the interval between the grooves is less than 200 nm.

  In the groove type sample target, the width and depth of the groove are preferably 1 nm or more and less than 30 μm, and more preferably 10 nm or more and less than 1 μm. According to said structure, it is easy to capture | acquire the energy of the laser beam of the ultraviolet region of several hundred nm order like a 337 nm nitrogen laser etc., and can obtain favorable ionization efficiency. In order to better ionize the measurement sample, it is more preferable that the interval between the grooves is 10 nm or more and less than 200 nm.

<Concavity having pore shape>
When the shape of the recess is a pore, the pore opens on the surface of the sample holding surface and extends in the thickness direction of the sample holding surface. The pore arrangement, shape, and angle with the sample holding surface may be regular or irregular, but in order to improve the function as a sample target for mass spectrometry, More preferably, it is regular.

  An example of such a sample holding surface is schematically shown in FIG. 4A is a perspective view showing a part of the sample holding surface of the sample target, and FIG. 4B is a cross-sectional view of the sample holding surface showing a cut surface taken along the broken line B in FIG. In FIG. 4, the sample holding surfaces shown in (a) and (b) indicate sample holding surfaces in the case where the arrangement and shape of the pores and the angle with the sample holding surface are regular. The sample holding surface is a surface of the sample holding surface that is irradiated with laser light, that is, a plurality of pores that are open on the upper surface of FIG. As shown in (b), the pores extend from the surface of the sample holding surface in the thickness direction of the sample holding surface and have a bottom.

  Moreover, the shape of the cross section when the pore is cut along a plane parallel to the sample holding surface is not particularly limited, and may be circular, elliptical, triangular, or quadrangular. Further, it may be a polygon such as a pentagon or a hexagon, or a shape obtained by slightly deforming these. Further, the shape of the cross section may be regular or irregular. That is, a single shape need not occupy all parts of the sample holding surface. However, in order to further improve the function as a sample target for mass spectrometry, the cross-sectional shape is preferably regular, that is, the same shape. Further, in order to further improve the function as a sample target for mass spectrometry, the shape of the cross section is preferably constant from the opening to the bottom of the pore, but may be slightly deformed.

  The pores only need to extend in the thickness direction from the surface of the sample holding surface, and the pores are preferably perpendicular to the surface of the sample holding surface, but have a slight inclination. It doesn't matter. Further, the angle between the pores and the surface of the sample holding surface may be different for each pore, but is preferably regular. That is, it is preferable that each pore extends in the same direction. This is preferable because the function as a sample target can be further improved. The pores preferably extend linearly from the opening to the bottom. Since the pores are linear, the laser light enters the inside of the pores, which is preferable because ionization efficiency is good.

  In addition, the pore has a width of the concave portion, that is, a pore diameter of 30 nm or more and less than 5 μm, and the depth of the concave portion / (interval of the concave portion−the width of the concave portion), that is, the depth of the pore / (pore cycle). -Pore diameter) is more preferably 2 or more and 50 or less. Thereby, the ionization performance can be further improved. Here, the pore diameter means the diameter when the cross section obtained by cutting the pore with a plane parallel to the sample holding surface is a circle, and means the size of the portion indicated by D in FIG. In addition, when the cross section obtained by cutting the pore with a plane parallel to the sample holding surface is not a circle, the diameter of the circle whose circumference is the circumference is the width of the recess, that is, the pore diameter. Further, the depth of the concave portion, that is, the pore depth means the length from the opening to the bottom of the pore, and means the size of the portion indicated by E in FIG. Further, the pore period means the interval between the concave portions, that is, the interval between the center lines of adjacent pores, and means the size of the portion indicated by C in FIG. In FIG. 4, the alternate long and short dash line is the center line of the pore. When the pore diameter, pore depth, and pore cycle are not uniform, these values are obtained using average values. The cross-sectional view shown in FIG. 4 is a cross-sectional view cut along a cross section where D is the largest. For example, when the cross section obtained by cutting the pore with a plane parallel to the sample holding surface is circular, it is a cross sectional view cut with a plane including the diameter.

  The pore period and the pore diameter may be about 1 nm to several tens of μm. However, in order to further improve the function as a sample target for mass spectrometry, the pore period is 30 nm or more and less than 5 μm. More preferably, it is more preferably 31 nm or more and 1 μm, particularly preferably 33 nm to 500 nm, and most preferably 34 nm to 300 nm. The pore diameter is more preferably 30 nm or more and less than 5 μm, further preferably 40 nm or more and 1 μm, particularly preferably 45 nm to 700 nm, and most preferably 50 nm to 500 nm. Thereby, ionization of the measurement sample in mass spectrometry can be performed satisfactorily.

  The pore period and pore diameter may be regular or irregular. However, in order to further improve the function as a sample target for mass spectrometry, it is preferable to be regular. That is, it is preferable that the pore period and the pore diameter are uniform. When the pore period and the pore diameter are regular, the ionization performance is more stable because there is little variation in the structure having the concave portion of the sample holding surface.

  The pore depth may be 1 nm or more and less than 30 μm, but more preferably about 30 nm or more and less than 5 μm. However, in order to further improve the function as a sample target for mass spectrometry, it is more preferably 30 nm to 2 μm, further preferably 50 nm to 1.5 μm, and particularly preferably 70 nm to 1 μm. 100 nm to 1 μm is most preferable. The pore depth may be regular or irregular. That is, the pore depth may vary or be uniform. However, in order to further improve the function as a sample target for mass spectrometry, the pore depth is preferably uniform. If the pore depth is uniform, the unevenness of the sample holding surface is small, so that the ionization performance is more stable.

  Further, the pore depth / (pore cycle-pore diameter) is preferably 2 to 50, more preferably 2.5 to 45, and more preferably 3 to 35. Is more preferably 3.5-30, and most preferably 4-25. Thereby, even in the case where a matrix is not used in mass spectrometry, ionization performance can be further improved, and high molecular weight substances can be favorably ionized.

  When the value of pore depth / (pore cycle−pore diameter) is 50 or less, the strength of the surface of the sample holding surface can be maintained, and ionization is performed because laser light enters the pores. It can be suitably performed. Moreover, it is preferable at the point which is excellent in ionization efficiency because the value of pore depth / (pore period-pore diameter) is 2 or more.

  When the pores have an irregular structure, the value of pore depth / (pore cycle−pore diameter) is an average value of the entire portion where the pores are arranged (porous portion). The value of pore depth / (pore period−pore diameter) is determined without considering partial large defects.

(I-2) Coating with metal and / or semiconductor In the sample holding surface of the sample target according to the present invention, the surface of the sample holding surface excluding a part of the inner surface of the recess is coated with metal and / or semiconductor. In addition, a portion not covered with metal and / or semiconductor is present on the inner surface of the recess. Thereby, ionization performance can be remarkably improved, and ionization of a substance having a molecular weight exceeding 30000 becomes possible.

  FIG. 2 is a diagram schematically showing coating of the surface of the sample holding surface with metal and / or semiconductor when porous alumina is used as a sample target as an example of the present invention. 2A shows a conventional sample holding surface in which the entire surface of the sample holding surface including the inner surface of the concave portion is covered with metal and / or semiconductor, and FIG. 2B shows the concave portion of the concave portion according to the present invention. The sample holding surface in which the part which is not coat | covered with the metal and / or semiconductor exists in an inner surface is shown. In FIG. 2, a portion indicated by 1 indicates a portion covered with a metal and / or a semiconductor. In (b), the surface excluding the inner surface of the concave portion of the sample holding surface and the upper portion including the upper end portion of the side wall of the inner surface of the concave portion are covered with metal and / or semiconductor. As shown in (b), the sample holding surface used in the present invention has at least a surface other than the inner surface of the recess covered with a metal and / or a semiconductor. Thus, by covering at least the surface other than the inner surface of the recess with metal and / or semiconductor, the conductivity can be increased at least on the surface other than the inner surface of the recess, and ionization performance can be improved. It is considered possible.

  The area of the inner surface of the recess that is not covered with metal and / or semiconductor is preferably 10% or more and 100% or less, and 20% or more and 100% or less with respect to the entire inner surface of the recess. It is more preferable that it is 30% or more and 100% or less. The ratio of the area of the portion not covered with metal and / or semiconductor to the area of the entire inner surface is within the above range, thereby significantly improving ionization performance and ionizing a high molecular weight substance having a molecular weight exceeding 30000. It becomes possible. In addition, when the ratio of the area of the part which is not coat | covered with a metal and / or a semiconductor with respect to the area of the whole inner surface is not uniform for every recessed part, it is set as these values using an average value.

  In addition, the part coat | covered with the metal and / or semiconductor and the part which is not coat | covered can be confirmed with a scanning electron microscope (SEM). That is, as shown in FIG. 1 which shows the result of observing a sample target whose upper part of the inner surface of the recess is covered with metal by SEM, the metal and semiconductor are secondary electrons compared to the material of the sample holding surface described later. High brightness is observed because it is easy to release. Therefore, by analyzing the SEM image, the ratio of the area of the portion not covered with the metal and / or semiconductor can be obtained.

  In addition, as a method for more clearly evaluating a portion covered with a metal and / or a semiconductor and a portion not covered, a case where the portion not covered is a dot (island) is not used. A method of confirming with a scanning electron microscope (SEM) after performing electrolytic deposition using the coated portion as an electrode can be suitably used. In such a method, a silver wire is fixed with a conductive paste on the surface of a sample target on which the surface of the sample holding surface excluding a part of the inner surface of the recess to be evaluated is coated with metal and / or semiconductor, and Ni plating is performed. Using a Ni plate as a counter electrode in liquid at room temperature, Ni is electrolytically deposited for 10 minutes under a constant voltage condition of -1V. When SEM observation of the cross section of the sample target after electrolytic deposition is performed, Ni is selectively electrolytically deposited only on the portion covered with the metal and / or semiconductor, so that it is coated with the metal and / or semiconductor. The part and the uncoated part can be more clearly evaluated.

  The position of the inner surface of the recess that is not covered with metal and / or semiconductor is not particularly limited, and includes, for example, the upper end portion of the inner wall of the recess as shown in FIG. It may be a portion other than the top and the bottom, may be the entire inner surface of the recess, may be a part of the inner sidewall of the recess, may be the entire inner sidewall of the recess, It may be only the bottom surface of the recess, or the covered part may be dot-like (island-like) and may be a part other than the dot-like (island-like), or the part that is not covered is a dot (Island shape).

  In the sample target according to the present invention, the portion of the inner surface of the recess that is covered with metal and / or semiconductor is the upper portion of the sidewall including the upper end portion of the sidewall of the recess, More preferably, the distance between the lower end of the covered portion is 0 or more on average and the width of the recess × 1.73 or less. Thereby, ionization performance can be further improved. Here, the width of the recess is as described in (I-1) above. The distance from the lower end of the covered portion is preferably 0 or more and the width of the recess × 1.73 or less on average, but is 0 or more and the width of the recess × 1.19 or less. More preferably, it is particularly preferably 0 or more and the width of the recess × 1 or less.

  Specific examples of the metal that can be used for coating the sample holding surface include, for example, group 1A (Li, Na, K, Rb, Cs, Fr) and group 2A (Be, Mg, Ca) of the periodic table of elements. , Sr, Ba, Ra), 3A group (Sc, Y), 4A group (Ti, Zr, Hf), 5A group (V, Nb, Ta), 6A group (Cr, Mo, W), 7A group (Mn) , Tc, Re), Group 8 (Fe, Ru, Os, Co, Rh, Ir, Ni, Pd, Pt), Group 1B (Cu, Ag, Au), Group 2B (Zn, Cd, Hg), Group 3B (Al), and lanthanoid series (La, Ce, Pr, Nd, Pm, Sm, Eu, Gd, Tb, Dy, Ho, Er, Tm, Yb, Lu), actinoid series (Ac, Th, Pa, U, Np, Pu, Am, Cm, Bk, Cf, Es, Fm, Md, No, Lr) Can. Of these, the metal is more preferably Au or Pt. Since Au and Pt are not easily oxidized, it is possible not only to improve ionization efficiency but also to prevent oxidation of the sample holding surface on which a plurality of concave portions are repeatedly formed.

  Further, the metal may be a single metal selected from the above metals, or an alloy composed of at least two or more selected from the above metals. Here, the alloy may be a metal in which two or more kinds of metals are mixed, and the existence form of the two or more kinds of mixed metals is not particularly limited. Examples of the presence form of the two or more kinds of mixed metals include a solid solution, an intermetallic compound, a state in which a solid solution and an intermetallic compound are mixed, and the like.

Specific examples of the semiconductor that can be used for coating the sample holding surface include Si, Ge, SiC, GaP, GaAs, InP, Si _1-X Ge _X (0 <X <1), _{SnO 2, ZnO, In 2 O} 3 and mixtures thereof, and carbon or the like. Among these, the semiconductor is more preferably SnO ₂ , ZnO, In ₂ O ₃ , ITO which is a mixture of SnO ₂ and In ₂ O ₃ , or the like. Since these substances are originally oxides and are not oxidized any more, the ionization performance is not lowered even when left in the air. In addition, although carbon has different physical properties depending on the bonding state of its atoms, it is classified here as a semiconductor. Since carbon is not easily oxidized in the air, the performance of ionization is not lowered even if it is left in the air.

  The surface of the sample holding surface may be coated with a mixture of at least two selected from the semiconductor and the metal.

  The thickness of the coated metal and / or semiconductor is not particularly limited as long as it does not impair the structure having the concave portion of the sample holding surface. Specifically, for example, it is preferably 1 nm or more and 200 nm or less. When the thickness of the metal and / or semiconductor does not exceed this upper limit, the structure of the sample holding surface is not impaired, and when the thickness is larger than the lower limit, efficient ionization is possible. Furthermore, the thickness of the metal and / or semiconductor is more preferably 1 nm or more and 150 nm or less, further preferably 5 nm or more and 100 nm or less, particularly preferably 10 nm or more and 80 nm or less, and 20 nm or more and 75 nm or less. Most preferably. Thereby, more efficient ionization becomes possible.

(I-3) Material of Sample Holding Surface The material of the sample holding surface is not particularly limited as long as it has the above shape, and examples thereof include resins such as synthetic polymers, ceramics, and the like. Even if it is a material which does not have electroconductivity, the efficiency of ionization can be improved by coat | covering with a metal and / or a semiconductor.

  Examples of the synthetic polymer include polyethylene, polypropylene, polyacrylic acid ester, polymethacrylic acid ester, polystyrene, polysiloxane, polystannoxane, polyamide, polyester, polyaniline, polypyrrole, polythiophene, polyurethane, polyethyl ether ketone, and poly-4-fluoride. A copolymer containing at least one of these; a mixture containing at least one of these; a graft polymer containing at least one of these; a block polymer containing at least one of these, and the like.

  The ceramics include alumina (aluminum oxide), magnesia, beryllia, zirconia (zirconium oxide), uranium oxide, thorium oxide, silica (quartz), holsterite, steatite, wollastonite, zircon, mullite, cordierite / Cordierite, spodumene, aluminum titanate, spinel apatite, barium titanate, ferrite, lithium niobate, silicon nitride (silicon nitride), sialon, aluminum nitride, boron nitride, titanium nitride, silicon carbide (silicon carbide), boron carbide , Titanium carbide, tungsten carbide, lanthanum boride, titanium boride, zirconium boride, cadmium sulfide, molybdenum sulfide, molybdenum silicide, diamond, single crystal sapphire, etc. That.

  As the material for the sample holding surface, for example, porous alumina can be suitably used. Porous alumina refers to an oxide film having many fine pores formed on the surface by anodizing aluminum or an alloy thereof in an electrolytic solution. By controlling the anodizing conditions, it is possible to produce regular porous alumina in which pores are regularly arranged over a wide range. In the regular porous alumina thus obtained, for example, as shown in FIG. 5, a large number of pores are arranged in one direction on a layer 102 of aluminum (or an alloy thereof) via a barrier layer 103. A porous alumina layer 101 is formed. Although the barrier layer is present in FIG. 5, the barrier layer may be removed.

  Thus, porous alumina has a plurality of regularly formed recesses, and can obtain a structure having a preferable pore depth / (pore cycle-pore diameter) in the sample target. It can use suitably for the sample target of invention. Furthermore, porous alumina can be suitably used in the present invention because the pore diameter, pore depth, and pore cycle can be controlled by changing the anodizing conditions.

  Porous alumina having a regular pore structure is disclosed in, for example, H. Masuda and M. Satoh, Jpn. J Appl. Phys., 35, pp. L126 (1996) in two steps. A method of performing oxidation, disclosed in JP-A-10-121292, after a substrate having a plurality of protrusions is applied to the surface of an aluminum plate to be anodized to form a desired pore period and an array of depressions, It can be obtained by a conventionally known method such as a method of anodizing the aluminum plate.

  Further, as the sample holding surface, a negative second structure in which the concave / convex structure of the first structure is transferred using the first structure having a concave / convex structure in which fine concave portions are repeatedly formed as a mold is prepared. In addition, a sample holding surface having a concavo-convex structure having the same shape as the concavo-convex structure of the first structure, which is obtained by transferring the concavo-convex structure using the negative second structure as a mold, is preferably used. it can. By using such a sample holding surface, a sample target having a surface on which a plurality of concave portions can be repeatedly formed, which can be used in the present invention, can be manufactured using various materials.

  The first structure having a concavo-convex structure in which fine concave portions are repeatedly formed is not particularly limited as long as it has a concavo-convex structure suitable for the sample target of the present invention. For example, porous alumina is used. It can be used suitably.

  The material of the sample holding surface, to which the concavo-convex structure is transferred using the negative second structure as a mold, is not particularly limited. For example, a material containing the resin, ceramics, or the like can be suitably used. .

(II) Sample target manufacturing method A sample target manufacturing method according to the present invention is a method for manufacturing the sample target, wherein the surface of the sample holding surface is coated with a metal and / or a semiconductor on the inner surface of the recess. A coating step of coating with a metal and / or semiconductor is included so that there is an unfinished portion.

(II-1) Coating step In the coating step, the surface of the sample holding surface is coated with a metal and / or semiconductor so that there is a portion not covered with metal and / or semiconductor on the inner surface of the recess. The method is not particularly limited as long as it can be used, and a conventionally known method can be used. Examples of such methods include sputtering, chemical vapor deposition (CVD), vacuum deposition, electroless plating, electroplating, coating, noble metal varnish, organometallic thin film, and sol-gel. it can.

  Among them, as a method for coating the inner surface of the recess so that there is a portion not coated with metal and / or semiconductor, physical vapor deposition is used and the sample holding surface is extended in the thickness direction. A method of depositing the metal and / or semiconductor can be suitably used from an angle inclined obliquely from the direction of the center line or the center plane of the recess. By using such a method, it is possible to simply perform coating so that a portion not covered with metal and / or semiconductor exists on the inner surface of the recess by simply changing the deposition angle.

  Here, the physical vapor deposition is not particularly limited. For example, sputtering such as ion beam sputtering, direct current sputtering, magnetron sputtering, and high frequency sputtering, thermal vapor deposition, electron beam vapor deposition, and vacuum vapor deposition are preferably used. Can do.

  In this step, the metal and / or semiconductor is vapor-deposited from the center line of the recess extending in the thickness direction from the surface of the sample holding surface or from an angle inclined obliquely from the direction of the center surface. FIG. 2 schematically shows the coating of the surface of the sample holding surface with metal and / or semiconductor when porous alumina is used as a sample target as an example of the present invention. In FIG. 2, (a) shows the surface of the conventional sample holding surface where the whole sample holding surface including the inner surface of a recessed part was coat | covered with the metal and / or the semiconductor, (b) shows the recessed part concerning this invention. The surface of the sample holding surface where the inner surface includes a portion not covered with metal and / or semiconductor is shown.

  In FIG. 2, the alternate long and short dash line indicates “the direction of the center line of the recess extending in the thickness direction from the surface of the sample holding surface”. This direction is the direction of the center line of the pore extending in the thickness direction from the surface of the sample holding surface when the recess is a pore, and from the surface of the sample holding surface when the recess is a groove. This is the direction of the center plane of the groove extending in the thickness direction.

  Moreover, the arrow in FIG. 2 shows the direction of vapor deposition, ie, the flight direction of the particles that adhere to the surface of the sample holding surface and coat the sample holding surface. As shown in FIG. 2A, conventionally, the metal and / or semiconductor is deposited from the direction parallel to the direction of the recess extending in the thickness direction from the surface of the sample holding surface. In contrast, in the sample target manufacturing method according to the present invention, as shown in (b), the center line of the recess extending in the thickness direction from the surface of the sample holding surface or obliquely from the direction of the center surface. The metal and / or semiconductor is deposited from an inclined angle. As a result, as shown in the figure, the metal and / or semiconductor does not enter the entire inside of the recess, and the inner surface of the recess of the sample holding surface is covered only with the metal and / or semiconductor. Therefore, the lower part of the recess where the metal and / or semiconductor does not enter remains as a part not covered with the metal and / or semiconductor. Therefore, it is possible to ionize a high molecular weight substance having a molecular weight exceeding 30000 and to significantly improve the ionization performance of the low molecular weight compound.

  Here, the angle inclined obliquely from the center line of the recess extending in the thickness direction from the surface of the sample holding surface or the direction of the center surface is an angle indicated by θ in FIG. It can also be said to be an angle formed by the direction of the center line or the center plane of the recess extending in the thickness direction from the surface of the film and the direction of vapor deposition. It is preferable that the angle θ tilted obliquely from the center line of the recess extending in the thickness direction from the surface of the sample holding surface or from the direction of the center surface is 30 ° or more and less than 90 °. This makes it possible to ionize a high molecular weight substance having a molecular weight exceeding 30000 and to significantly improve the ionization performance of a low molecular weight compound. Further, the angle θ is more preferably 40 ° or more and less than 89 °, further preferably 45 ° or more and less than 88 °, and particularly preferably 60 ° or more and less than 85 °. Thereby, ionization performance can be further improved.

  FIG. 10 shows, for example, when Pt is used as the metal and Pt is vapor-deposited on the sample holding surface made of porous alumina, the center line or the center plane of the recess extending in the thickness direction from the surface of the sample holding surface. It is a schematic diagram which shows the angle | corner which the direction and the direction of vapor deposition make, and the part coat | covered with Pt.

  As shown in FIG. 10, the portion covered with the metal and / or semiconductor on the inner surface of the recess can be defined by the angle θ and the width of the recess. That is, the portion of the inner surface of the recess covered with metal and / or semiconductor is the upper portion of the side wall including the upper end of the side wall of the recess, and the upper end of the side wall and the lower end of the covered portion Is represented by Tan (90 ° −θ) × D. Here, D refers to the width of the recess and is as described in (I-1) above.

  2 and 10, the direction in which the metal and / or semiconductor is vapor-deposited includes the center line of the recess extending in the thickness direction from the surface of the sample holding surface or the direction of the central surface and the direction of vapor deposition. Only one direction where the angle formed is θ is shown, but the sample holding surface is rotated while maintaining θ at the same angle, so that only the desired range is covered in all directions of the inner surface of the recess. Can do.

  When the concave portion is a groove, the sample holding surface may be rotated while avoiding a range where the vapor deposition direction and the longitudinal direction of the groove are the same.

(II-2) Method for Producing Sample Holding Surface As a method for producing a sample holding surface for coating a metal and / or semiconductor in the coating step, a method capable of producing the sample holding surface described in (I) above. If it is, it will not specifically limit. Among these, as a method for producing such a sample holding surface, for example, a method using porous alumina, a first structure having a concavo-convex structure in which fine concave portions are repeatedly formed is used as a mold, and the first structure used as a mold is used. A method for manufacturing a sample holding surface made of another material having the same concavo-convex structure, a method using lithography, and the like can be suitably used.

<Method using porous alumina>
Here, the porous alumina may be produced by anodizing aluminum or an alloy thereof, or commercially available porous alumina may be used.

  The method for producing porous alumina is not particularly limited, and any method may be used. Moreover, a conventionally well-known method can be used suitably. In general, aluminum or an alloy thereof is preferably polished and anodized in an electrolytic solution. The electrolytic solution may be acidic or alkaline, but is preferably sulfuric acid, oxalic acid, phosphoric acid, or the like. In addition, in order to obtain a sample holding surface having a desired pore diameter, pore depth, and pore cycle, the anodizing voltage, anodizing time, the type and concentration of the electrolytic solution, temperature conditions, and the like may be appropriately selected. In addition, the method of polishing aluminum or an alloy thereof before anodic oxidation is not particularly limited. For example, a method of electrolytic polishing in a mixed solution of perchloric acid and ethanol, a mixed solution of phosphoric acid and sulfuric acid, or the like. And a method of mechanically polishing the surface. The porous alumina obtained by anodic oxidation may be subjected to an enlargement treatment of the pore diameter by an etching treatment using a phosphoric acid aqueous solution, a sulfuric acid aqueous solution or the like.

  In order to produce regular porous alumina, as described above, for example, it is disclosed in H. Masuda and M. Satoh, Jpn. J Appl. Phys., 35, pp. L126 (1996). A method of performing anodization in two stages, disclosed in JP-A-10-121292, a substrate (mold) having a plurality of protrusions is applied to the surface of an aluminum plate to be anodized to obtain desired pores A method of anodizing the aluminum plate after forming the recesses of the period and arrangement can be suitably used.

<Nanoimprint method>
As a method of manufacturing a sample holding surface made of another material having the same concavo-convex structure as the first structure using the first structure having the concavo-convex structure in which fine concave portions are repeatedly formed as a mold. Any method may be used as long as it is a “nanoimprint” method in which a microstructure is used as a template and the structure is transferred to another substance. In recent years, in the field of nanotechnology, in order to fabricate DNA chips, semiconductor devices, minute containers for chemical reactions, etc., a microstructure manufactured in units of 1 nm to several tens of μm is used as a template. Various “nanoimprint” methods for transferring the structure to another substance have been developed, and these conventionally known methods can be suitably used.

  Further, by using such a nanoimprint method, a sample holding surface having the same structure as the first structure having an uneven structure suitable for the sample holding surface can be manufactured with a desired material. Therefore, these manufacturing methods are also included in the present invention.

  That is, when the nanoimprint method is used for manufacturing the sample holding surface, the sample target manufacturing method according to the present invention repeats the fine recesses before the step of coating the surface of the sample holding surface with metal and / or semiconductor. Using the formed first structure having a concavo-convex structure as a mold to produce a negative second structure in which the concavo-convex structure of the first structure is transferred, and a negative type first structure obtained in the step There may be included a step of transferring the concavo-convex structure using two structures as a mold to obtain a sample holding surface having a concavo-convex structure having the same shape as the concavo-convex structure of the first structure on the surface.

  The first structure is not particularly limited as long as it has a concavo-convex structure suitable for the sample target of the present invention. For example, porous alumina can be suitably used.

  Examples of such nanoimprinting methods include the method described in T. Yanagishita, K. Nishino, H. Masuda: Jpn. J. Appl. Phys., Vol. 45, No. 30 (2006). In this method, a negative second structure in which the uneven structure of porous alumina is transferred to Ni is produced using porous alumina as a mold. Using the obtained negative second structure made of Ni as a mold, a sample holding surface made of a polymer having the uneven structure on its surface is manufactured.

  An example of the above method will be described below more specifically based on FIG. FIG. 7 is a diagram schematically illustrating a sample target manufacturing method including a sample holding surface manufacturing step by a nanoimprint method. First, in order to form an electrode for electrolytic deposition of Ni, a metal film of 5 nm to 200 nm is formed on the surface of porous alumina by sputtering, and Ni is electrolytically deposited ((i) in FIG. 7). Here, the metal film is not particularly limited as long as Ni can be electrolytically deposited. For example, a Pt film, an Au film, a Ta film, a Pd film, or the like can be suitably used. . After electrolytic deposition, the porous alumina is dissolved and removed to obtain a negative second structure in which the uneven structure of the porous alumina is transferred to Ni ((ii) in FIG. 7). The negative second structure made of Ni is pretreated with a release agent as necessary, and then a photocurable resin monomer is poured onto the second structure, for example, a glass substrate or the like. With the substrate attached, the monomer is polymerized under UV irradiation. The negative second structure made of Ni is mechanically removed ((iii) in FIG. 7) to obtain a sample holding surface ((iv) in FIG. 7) made of a polymer having a concavo-convex structure of porous alumina on the surface. . (V) and (vi) in FIG. 7 schematically show the subsequent coating step.

  The material used as a negative structure for transferring the concavo-convex structure of the first structure having a concavo-convex structure in which fine concave portions are repeatedly formed used in such a method is not limited to Ni, for example, Group 1A (Li, Na, K, Rb, Cs, Fr), Group 2A (Be, Mg, Ca, Sr, Ba, Ra), Group 3A (Sc, Y), Group 4A (Ti, Zr) , Hf), Group 5A (V, Nb, Ta), Group 6A (Cr, Mo, W), Group 7A (Mn, Tc, Re), Group 8 (Fe, Ru, Os, Co, Rh, Ir, Ni) , Pd, Pt), 1B group (Cu, Ag, Au), 2B group (Zn, Cd, Hg), 3B group (Al), and lanthanoid series (La, Ce, Pr, Nd, Pm, Sm, Eu, Gd, Tb, Dy, Ho, Er, Tm, Yb, Lu), Actinoy Sequence (Ac, Th, Pa, U, Np, Pu, Am, Cm, Bk, Cf, Es, Fm, Md, No, Lr) metal may be used, such as.

  In addition, the polymer that transfers the uneven structure to the surface using the negative second structure as a mold is not particularly limited, and polyethylene, polypropylene, polyacrylate, polymethacrylate, polystyrene, Polysiloxane, polystannoxane, polyamide, polyester, polyaniline, polypyrrole, polythiophene, polyurethane, polyethyletherketone, poly-4-fluoroethylene; copolymer containing at least one of these; mixture containing at least one of these; A graft polymer containing at least one; a block polymer containing at least one of these can be used.

  The solvent used for dissolving and removing the porous alumina is not particularly limited as long as it dissolves aluminum and alumina, and examples thereof include an aqueous sodium hydroxide solution and an aqueous potassium hydroxide solution.

  In the nanoimprint method described above, the method of transferring the concavo-convex structure of the first structure having the concavo-convex structure in which fine concave portions are repeatedly formed in the resin was shown. However, the concavo-convex structure of the first structure was transferred by the nanoimprint method. The material of the sample holding surface produced in this way is not limited to resin, and may be ceramics, for example.

  Moreover, as this ceramic, the same ceramics as described in the above (I-3) can be suitably used.

<Lithography>
As a method using lithography, there are a photolithography method, an electron beam lithography method, an ion beam lithography method, a nanoimprint lithography method, a dip pen nanolithography method, and the like. Of these lithography methods, it is more preferable to use an electron beam lithography method. If the electron beam lithography method is used, since the size of the writing shape is not limited by the wavelength of light as in general optical lithography, finer writing can be performed. The recesses can be repeatedly formed such that the distance between is 1 nm or more and less than 30 μm.

  In the electron beam lithography method, a device design drawing is baked on a metal plate called a mask, and a part of the mask transmits light, and the other part does not transmit light. Then, when light is applied to the processed design drawing and the light is focused by a lens, the design drawing pattern is reduced and projected. Here, a photosensitizer is applied to the material that becomes the base of the device in advance, and when the projection is reduced and projected onto the base, the pattern of the design drawing is printed there.

  The photosensitive agent applied to the substrate is called a resist. The resist uses molecules that cause a photoreaction, such as solidifying when exposed to light or becoming insoluble in a polymerized solution. When the material of the substrate on which the pattern has been baked is put into a solution for dissolving it, only the portion where the resist that has been exposed to light is hardened is melted, and the other portions can be melted. By further etching using the resist pattern formed in this way, it becomes possible to perform fine processing on the substrate. In the electron beam lithography method, an electron beam drawing apparatus is generally used. The accuracy of creating the fine structure greatly depends on the performance of the electron beam drawing apparatus.

  As described above, when the sample target according to the present invention is manufactured, if the lithography technique is used, the recesses can be repeatedly formed on the sample holding surface so that the interval between the recesses is 1 nm or more and less than 30 μm.

  Therefore, in the sample target manufacturing method of the present invention, before the step of coating the surface of the sample holding surface with metal and / or semiconductor, the space between the recesses is 1 nm or more on the surface of the substrate using the lithography technique. A step of forming a sample holding surface on the surface may be included by repeatedly forming the recess so as to be less than 30 μm.

(II-3) Step of Enlarging the Diameter of the Concave The method for producing a sample target according to the present invention further includes etching the portion of the inner surface of the concave that is not covered with metal and / or semiconductor, A step of expanding the diameter may be included.

  Such a step is preferably included in the method for manufacturing a sample target according to the present invention if the inner surface of the recess is not made of a metal and / or semiconductor and is made of a material that can be removed by etching. Thereby, although the reason is not clear, the ionization performance can be further improved.

  FIG. 6 is a diagram schematically showing the sample holding surface obtained by the step of enlarging the diameter of the concave portion of the portion not covered with the metal and / or semiconductor by the step of enlarging the diameter of the concave portion. FIG. 6 shows a sample holding surface in which the diameter of a portion not covered with metal and / or semiconductor is enlarged when porous alumina is used as the sample holding surface.

(III) Use of the present invention (mass spectrometer)
The sample target of the present invention is used as a sample stage for placing a sample to be measured in mass spectrometry of various substances such as biopolymers, endocrine disrupting substances, synthetic polymers, and metal complexes. be able to. The sample target is useful because it can efficiently and stably ionize the sample, particularly when used in laser desorption ionization mass spectrometry.

  Therefore, a mass spectrometer using the above-described sample target of the present invention is also included in the category of the present invention. The sample target can efficiently and stably ionize the sample, particularly when used in a laser desorption ionization mass spectrometer. Therefore, more specifically, the mass spectrometer of the present invention is a laser desorption ionization mass spectrometer that ionizes the sample to be measured by irradiating the sample with laser light and measures the molecular weight of the sample. preferable.

  In the laser desorption ionization mass spectrometer, the sample to be measured is placed on the sample target and used, so that the sample can be ionized well when irradiated with laser light. It can be carried out.

  The present invention will be described more specifically based on examples, but the present invention is not limited to this.

[Example 1]
An aluminum plate having a purity of 99.99% was subjected to electropolishing treatment in a perchloric acid / ethanol mixed solution (volume ratio 1: 4). A SiC mold having a structure in which protrusions were regularly arranged with a period of 200 nm was pressed onto the surface of the mirror-finished aluminum plate to form depressions with a fine uneven pattern on the surface. The textured aluminum plate is anodized in a 0.5 M aqueous phosphoric acid solution at a bath temperature of 17 ° C. under a direct current of 80 V for 15 minutes to form anodized porous alumina having a pore depth of 500 nm. did. Thereafter, the obtained anodized porous alumina was immersed in a 10% by weight phosphoric acid aqueous solution for 10 minutes, and subjected to pore size expansion treatment to adjust the pore size to 100 nm. The obtained anodized porous alumina was used as a sample holding surface. While the angle formed by the direction of the center line of the pore extending in the thickness direction from the surface of the sample holding surface and the direction of vapor deposition is 80 °, while rotating the sample holding surface using an ion beam sputtering apparatus, By coating Pt with 50 nm, the surface of the sample holding surface excluding a part of the inner surface of the recess is coated with Pt having a pore period of 200 nm, and the inner surface of the recess is coated with metal and / or semiconductor. A sample holding surface with a non-existing portion was obtained. Pore depth / (pore cycle−pore diameter) was 5.

  Next, mass spectrometry by laser desorption ionization was performed using the obtained sample target. 0.2 μL of 50% methanol solution of 5 μM peptide (angiotensin-I, molecular weight 1295.7) was dropped on the sample target, and after drying, a time-of-flight mass spectrometer Voyager DE-Pro (manufactured by Applied Biosystems) was used. Then, mass spectrometry by laser desorption ionization was performed in the linear mode.

FIG. 9 shows a mass spectrum obtained by irradiating a nitrogen laser (wavelength 337 nm, pulse width 10 nsec, repetition frequency 20 hertz) at a laser intensity of 35 mJ / cm ² . What is observed is the protonated ion [M + H] ⁺ .

In [Comparative Example 1] described later, the angle formed by the direction of the pores extending in the thickness direction from the surface of the sample holding surface (that is, the center line of the pores) and the direction of vapor deposition is 0 °, and Pt contrast from 50nm protonated ions when coated [M + ^{H] +,} was 204 counts, in the present embodiment, protonated ion [M + ^{H] +} is a 5203 count was strength of at least 25 times.

  From the results of this example, by depositing metal from an angle inclined from the direction of the recess extending in the thickness direction from the surface of the sample holding surface, the center line of the recess of the sample holding surface as in the past It can be seen that the ionization performance is remarkably improved as compared with the case where the metal is deposited from the direction parallel to.

[Example 2]
A sample target was produced in the same manner as in Example 1 except that the angle formed by the direction of the center line of the pore extending in the thickness direction from the surface of the sample holding surface and the direction of vapor deposition was 60 °.

  Next, using the obtained sample target, mass spectrometry by laser desorption ionization was performed in the same manner as in Example 1.

In [Comparative Example 1] described later, the angle formed by the direction of the pores extending in the thickness direction from the surface of the sample holding surface (that is, the center line of the pores) and the direction of vapor deposition is 0 °, and Pt contrast from 50nm protonated ions when coated [M + ^{H] +,} was 204 counts, in the present embodiment, protonated ion [M + ^{H] +} is a 1500 count was strength of at least 7 times.

[Comparative Example 1]
In the same manner as in Example 1, anodized porous alumina having a pore period of 200 nm, a pore diameter of 100 nm, and a pore depth of 500 nm was prepared and used for the sample holding surface. Using an ion beam sputtering apparatus, the sample is held by setting the angle formed by the direction of the pores extending in the thickness direction from the surface of the sample holding surface (that is, the center line of the pores) and the direction of vapor deposition to 0 °. While rotating the surface, 50 nm of Pt was coated to obtain a sample holding surface in which the entire surface of the sample holding surface including the inner surface of the recess was coated with Pt having a pore period of 200 nm. Pore depth / (pore cycle−pore diameter) was 5.

  Next, using the obtained sample target, the same measurement object as in Example 1 was used, and mass spectrometry by laser desorption ionization was performed under the same conditions.

FIG. 8 shows the obtained mass spectrum. What is observed is the protonated ion [M + H] ⁺ . The protonated ion [M + H] ⁺ was 204 counts.

Example 3
An aluminum plate having a purity of 99.99% was subjected to electropolishing treatment in a perchloric acid / ethanol mixed solution (volume ratio 1: 4). A SiC mold having a structure in which protrusions were regularly arranged with a period of 500 nm was pressed against the surface of the mirror-finished aluminum plate to form a recess with a fine uneven pattern on the surface. The textured aluminum plate is anodized in 0.1M phosphoric acid aqueous solution at a bath temperature of 0 ° C. under a direct current of 200 V for 4 minutes to form anodized porous alumina having a pore depth of 500 nm. did. Thereafter, the obtained anodized porous alumina was immersed in a 10% by weight phosphoric acid aqueous solution for 10 minutes, and subjected to pore size expansion treatment to adjust the pore size to 100 nm. The obtained anodized porous alumina was used as a sample holding surface. Sample holding is performed using an ion beam sputtering apparatus with the angle formed by the direction of pores extending in the thickness direction from the surface of the sample holding surface (that is, the center line of the pores) and the direction of vapor deposition being 80 °. By coating Pt with 50 nm while rotating the surface, the surface of the sample holding surface excluding a part of the inner surface of the recess is coated with Pt, and the inner surface of the recess is coated with metal and / or semiconductor. A sample holding surface with a portion that was not formed was obtained.

  Thereafter, the bottom of the pores was dissolved by etching the obtained sample holding surface in a mixed solution of chromic phosphate.

  Next, mass spectrometry by laser desorption ionization was performed using the obtained sample target. Bovine serum albumin having a molecular weight of 66000 was dissolved in 50% methanol to a concentration of 5 μM. 0.5 μL of the obtained bovine serum albumin methanol solution was supported on a sample target and linearized by a nitrogen laser 337 nm using a time-of-flight mass spectrometer Voyager DE-Pro (Applied Biosystems). Mass spectrometry by laser desorption ionization was performed in the mode.

  FIG. 11 b shows the obtained mass spectrum. As shown in FIG. 11b, ions of m / z 66000 could be detected.

[Comparative Example 2]
A sample target was produced in the same manner as in Example 3 except that the angle formed by the direction of the center line of the pore extending in the thickness direction from the surface of the sample holding surface and the direction of vapor deposition was 0 °.

  Next, mass spectrometry by laser desorption ionization was performed in the same manner as Example 3 using the obtained sample target.

  The mass spectrum obtained is shown in FIG. As shown in FIG. 11a, ions of m / z 66000 could not be detected.

Example 4
An aluminum plate having a purity of 99.99% was subjected to electropolishing treatment in a perchloric acid / ethanol mixed solution (volume ratio 1: 4). A SiC mold having a structure in which protrusions were regularly arranged with a period of 200 nm was pressed onto the surface of the mirror-finished aluminum plate to form depressions with a fine uneven pattern on the surface. This textured aluminum plate is anodized in 0.5 M phosphoric acid aqueous solution at a bath temperature of 17 ° C. under a direct current of 80 V for 11 minutes to form anodized porous alumina having a pore depth of 500 nm. did. Thereafter, the obtained anodized porous alumina was immersed in a 10% by weight phosphoric acid aqueous solution for 10 minutes, and subjected to pore size expansion treatment to adjust the pore size to 100 nm.

  Anodized porous alumina that had been subjected to pore diameter expansion treatment was coated with 50 nm of Pt using a sputtering apparatus, and then Ni electrolytic deposition was performed. Thereafter, the negative anodic porous alumina as a template was dissolved and removed, thereby obtaining a negative second structure made of Ni having a regular protrusion arrangement on the surface. A stock solution of a photocurable resin PAK-02A (manufactured by Toyo Gosei Co., Ltd.) was dropped onto the prepared second structure, and the resin was cured by irradiating with ultraviolet rays. After the photocurable resin was completely polymerized and solidified, the mold was peeled from the polymer layer to obtain a polymer hole array.

  Sample holding is performed using an ion beam sputtering apparatus with the angle formed by the direction of pores extending in the thickness direction from the surface of the sample holding surface (that is, the center line of the pores) and the direction of vapor deposition being 80 °. By coating Pt with 50 nm while rotating the surface, the surface of the sample holding surface excluding a part of the inner surface of the recess is coated with Pt, and the inner surface of the recess is coated with metal and / or semiconductor. A sample holding surface with a portion that was not formed was obtained. Pore depth / (pore cycle−pore diameter) was 5.

Next, mass spectrometry by laser desorption ionization was performed using the obtained sample target. 0.2 μL of 50% methanol solution of 5 μM peptide (angiotensin-I, molecular weight 1295.7) was dropped on the sample target, and after drying, a time-of-flight mass spectrometer Voyager DE-Pro (manufactured by Applied Biosystems) was used. Then, a nitrogen laser (wavelength: 337 nm, pulse width: 10 nsec, repetition frequency: 20 Hz) was irradiated at a laser intensity of 35 mJ / cm ^{2, and} mass spectrometry was performed in a linear mode by a laser desorption ionization method. The proton-added ion [M + H] ⁺ was 2719 counts.

Example 5
An aluminum plate having a purity of 99.99% was subjected to electropolishing treatment in a perchloric acid / ethanol mixed solution (volume ratio 1: 4). A SiC mold having a structure in which protrusions were regularly arranged with a period of 200 nm was pressed onto the surface of the mirror-finished aluminum plate to form depressions with a fine uneven pattern on the surface. The textured aluminum plate is anodized in a 0.5 M aqueous phosphoric acid solution at a bath temperature of 17 ° C. under a direct current of 80 V for 15 minutes to form anodized porous alumina having a pore depth of 500 nm. did. The obtained anodized porous alumina was used as a sample holding surface. Sample holding is performed using an ion beam sputtering apparatus with the angle formed by the direction of pores extending in the thickness direction from the surface of the sample holding surface (that is, the center line of the pores) and the direction of vapor deposition being 80 °. By coating Pt with 50 nm while rotating the surface, the surface of the sample holding surface excluding a part of the inner surface of the recess is coated with Pt having a pore period of 200 nm, and the inner surface of the recess is made of metal and A sample holding surface having a portion not covered with semiconductor was obtained.

  Thereafter, the bottom of the pores was dissolved by etching the obtained sample holding surface in a mixed solution of chromic phosphate.

Next, mass spectrometry by laser desorption ionization was performed using the obtained sample target. 0.2 μL of 50% methanol solution of 5 μM peptide (angiotensin-I, molecular weight 1295.7) was dropped on the sample target, and after drying, a time-of-flight mass spectrometer Voyager DE-Pro (manufactured by Applied Biosystems) was used. Then, a nitrogen laser (wavelength: 337 nm, pulse width: 10 nsec, repetition frequency: 20 Hz) was irradiated at a laser intensity of 35 mJ / cm ^{2, and} mass spectrometry was performed in a linear mode by a laser desorption ionization method. Protonated ion [M + H] ⁺ was 6416 counts.

  According to the sample target of the present invention, in the laser desorption ionization mass spectrometry, it is possible to realize excellent ionization performance and high-molecular-weight substance ionization even when a matrix is not used.

  Laser desorption ionization mass spectrometry is currently used in a wide range of fields as mass spectrometry for biopolymers, endocrine disruptors, synthetic polymers, metal complexes, and the like. Since the sample target of the present invention is an effective material for performing this laser desorption ionization mass spectrometry more accurately and stably, it can be said that the applicability of the present invention is high.

It is a figure which shows the result of having observed the sample target by which the upper part of the inner surface of the recessed part was coat | covered with the metal by SEM. It is a figure which shows typically the coating | cover of the surface of a sample holding surface by a metal and / or a semiconductor at the time of using a porous alumina for a sample target. It is a schematic diagram which shows the shape of the groove | channel of a sample target in case a recessed part is a groove | channel. It is a schematic diagram which shows the shape of the pore of a sample target in case a recessed part is a pore. It is a sectional view showing the prior art and schematically showing regular porous alumina. It is a figure which shows typically the sample holding surface obtained by the process of expanding the diameter of the recessed part of the part which is not coat | covered with a metal and / or a semiconductor. It is a figure which shows typically the manufacturing method of the sample target including the process of a nanoimprint method. 3 is a mass spectrum obtained by performing mass spectrometric measurement of angiotensin-I using the sample target prepared in Comparative Example 1. 2 is a mass spectrum obtained by performing mass spectrometric measurement of angiotensin-I using the sample target prepared in Example 1. When depositing Pt on a sample holding surface made of porous alumina, the angle θ and Pt formed by the center line of the recess extending in the thickness direction from the surface of the sample holding surface or the direction of the center surface and the direction of vapor deposition. It is a schematic diagram which shows the part covered. It is the mass spectrum obtained by performing mass spectrometry measurement of bovine serum albumin using the sample target produced in Example 3 and Comparative Example 2.

Explanation of symbols

DESCRIPTION OF SYMBOLS 1 Metal and / or semiconductor 101 Porous alumina layer 102 Aluminum (or its alloy) layer 103 Barrier layer

Claims (20)

When performing mass spectrometry by ionizing a sample by laser light irradiation, it is used to hold the sample, and the surface on which a plurality of recesses that open to the surface that receives laser light irradiation are repeatedly formed is used as the sample holding surface. A sample target comprising:
The interval between adjacent recesses is 1 nm or more and less than 30 μm,
The surface of the sample holding surface excluding a part of the inner surface of the recess is coated with metal and / or semiconductor,
A sample target characterized in that a portion not covered with a metal and / or a semiconductor exists on the inner surface of the recess.   The portion of the inner surface of the recess that is covered with metal and / or semiconductor is the upper portion of the sidewall including the upper end of the sidewall of the recess, and is between the upper end of the sidewall and the lower end of the covered portion. The sample target according to claim 1, wherein the distance is an average of 0 or more and the width of the concave portion × 1.73 or less.   The sample target according to claim 1, wherein the plurality of concave portions are regularly and repeatedly formed.   The sample target according to claim 1, wherein the recess is a groove or a pore.   The sample target according to any one of claims 1 to 4, wherein the sample holding surface is made of porous alumina.   The sample holding surface is prepared using a first structure having a concavo-convex structure in which fine concave portions are repeatedly formed as a mold to produce a negative second structure in which the concavo-convex structure of the first structure is transferred, The sample holding surface having a concavo-convex structure having the same shape as the concavo-convex structure of the first structure, wherein the concavo-convex structure is transferred using a negative second structure as a mold. The sample target according to any one of 1 to 4.   The sample target according to claim 6, wherein the first structure is porous alumina.   The sample target according to claim 6 or 7, wherein the sample holding surface contains resin or ceramics.   The sample target according to claim 1, wherein the metal is platinum (Pt) or gold (Au). The _{semiconductor, Si, Ge, SiC, GaP} , GaAs, InP, Si 1-X Ge X (0 <X <1), SnO 2, ZnO, In 2 O 3, or these two or more thereof, Or it is carbon, The sample target of any one of Claims 1-9 characterized by the above-mentioned. When performing mass spectrometry by ionizing a sample by laser light irradiation, it is used to hold the sample, and the surface on which a plurality of recesses that open to the surface that receives laser light irradiation are repeatedly formed is used as the sample holding surface. A sample target manufacturing method comprising:
A sample target comprising a coating step of coating the surface of the sample holding surface with a metal and / or semiconductor so that a portion not coated with the metal and / or semiconductor exists on the inner surface of the recess. Production method.   The coating step uses physical vapor deposition, and deposits the metal and / or semiconductor from the center line of the recess extending in the thickness direction from the surface of the sample holding surface or from an angle inclined from the direction of the central surface. The method of manufacturing a sample target according to claim 11.   The method of manufacturing a sample target according to claim 12, wherein the physical vapor deposition is sputtering, thermal vapor deposition, electron beam vapor deposition, or vacuum vapor deposition.   12. The method according to claim 11, further comprising a step of, after the covering step, etching a portion of the inner surface of the recess that is not covered with metal and / or semiconductor to increase the diameter of the recess of the portion. The method for producing a sample target according to any one of ˜13.   The method for producing a sample target according to any one of claims 11 to 14, wherein porous alumina is used as a sample holding surface. Before the step of coating the surface of the sample holding surface with metal and / or semiconductor,
Using a first structure having a concavo-convex structure in which fine concave portions are repeatedly formed as a mold to produce a negative second structure in which the concavo-convex structure of the first structure is transferred;
Using the negative second structure obtained in this step as a mold, the concavo-convex structure is transferred to obtain a sample holding surface having a concavo-convex structure having the same shape as the concavo-convex structure of the first structure on the surface. The method of manufacturing a sample target according to claim 11, further comprising a step.   The method of manufacturing a sample target according to claim 16, wherein the first structure is porous alumina.   Prior to the step of coating the surface of the sample holding surface with metal and / or semiconductor, the concave portions are repeatedly formed on the surface of the substrate using a lithography technique so that the interval between the concave portions is 1 nm or more and less than 30 μm. The method of manufacturing a sample target according to any one of claims 11 to 14, further comprising a step of forming a sample holding surface on the surface.   A mass spectrometer using the sample target according to claim 1.   The mass spectrometer according to claim 19, wherein the mass spectrometer is a laser desorption / ionization mass spectrometer that irradiates a sample to be measured with laser light to ionize the sample and measure its molecular weight.