[Document Name] Description [Title of Invention] METHOD, DEVICE, AND PROGRAM FOR ANALYZING INTER-MOLECULAR INTERACTION [Technical Field] [0001] The present invention relates to a method, a device, and a program for analyzing inter-molecular interaction which analyze inter-molecular interaction. [Background Art] [0002] In recent years, with respect to the therapy for highly intractable diseases such as cancer, it has been active to find out the mechanism of the biological molecule function and to develop an antibody drug and the like blocking protein used for information transmission of cancer growth. The antibody used for an antibody drug is a multi-functional molecule having plural functions within the molecule. Therefore, respective evaluations of the function as the protein related to the action mechanism of an antibody drug and to the onset of action as well as structure/physical and chemical property generating the function become key items for securing effectiveness and safety of an antibody drug. [0003] A technology capable of evaluating these effectiveness and safety has been proposed in Patent Literature 1 for example. In Patent Literature 1, there is described an optical system biosensor that applies the surface plasmon resonance (SPR) phenomenon. In Patent Literature 1, it is described that, out of two molecules interacting with each other, one molecule called ligand in general is fixed to the sensor surface, and a specimen liquid including another molecule called analyte is added onto the sensor. Also, in Patent Literature 1, it is described that, by subsequently adding a test reagent not including an analyte such as a buffer onto the sensor, dissociation of the analyte having been bonded to the ligand can be observed. Further, in Patent Literature 1, it is described that a reaction rate constant and the like can be grasped from real-time change of the signal strength in accordance with the bonding/dissociation reaction of the ligand and the analyte on the sensor surface. [Citation List] [Patent Literature] [0004] [Patent Literature 1]

Description of U.S. Patent No. 9,316,636 [Summary of Invention] [Technical Problem] [0005] However, since the technology described in Patent Literature 1 uses a gold membrane for the sensor, there is a circumstance that biological matter and chemical matter are liable to be adsorbed. Therefore, according to the technology described in Patent Literature 1, it is required to remove other than the analyte to the maximum.

On the other hand, at the scene of development of drug development at present, there is a need to evaluate how an antibody drug effectively works actually in multiple components within a human being body, within the blood, on the cell, and so on, and whether a side effect is developed. That is to say, at the scene of drug development, there is a need to analyze inter-molecular interaction in a multiple component system including not only a first molecule and a second molecule which are the evaluation objects of inter-molecular interaction but also a third molecule. However, the technology described in Patent Literature 1 has the circumstance described above and multiple components cannot be included, and therefore there is a concern of not capable of responding to such needs.

[0006]

The object of the present invention is to provide a method, a device, and a program for analyzing inter-molecular interaction which can analyze not only inter-molecular interaction between a first molecule and a second molecule which is general as inter-molecular interaction but also an influence of a third molecule on inter-molecular interaction between these two molecules. [Solution to Problem]

[0007] A method for analyzing inter-molecular interaction related to the present invention includes a first step of fixing a first molecule to the surface of a magnetic sensor. The method for analyzing inter-molecular interaction related to the present invention includes a second step of bonding the first molecule and a second molecule to each other, the second molecule being labeled by a magnetic particle, and obtaining temporal change of a first magnetic signal. The method for analyzing inter-molecular interaction related to the present invention includes a third step of adding a third molecule other than the first molecule and the second molecule at prescribed timing after starting bonding the first molecule and the second molecule to each other and obtaining temporal change of a second magnetic signal. The method for analyzing inter-molecular interaction related to the present invention includes a fourth step of comparing a tendency of the temporal change of the first magnetic signal and a tendency of the temporal change of the second magnetic signal to each other, analyzing that the third molecule affects inter-molecular interaction between the first molecule and the second molecule when the tendencies of the temporal change are different from each other, and analyzing that the third molecule does not affect inter-molecular interaction when the tendencies of the temporal change are the same with each other. [Advantageous Effects of Invention]

[0008] According to the present invention, it is possible to provide a method, a device, and a program for analyzing inter- molecular interaction which can analyze not only an inter- molecular interaction between a first molecule and a second molecule which is general as inter-molecular interaction but also an influence of a third molecule on inter-molecular interaction between these two molecules.

Problems, configurations, and effects other than those described above will be clarified by explanation of embodiments described below.

[Brief Description of Drawings]

[0009]

[FIG. 1] FIG. 1 is a graphic chart showing a flow of a method for analyzing inter-molecular interaction related to an embodiment and contents in each step.

[FIG. 2] FIG. 2 is a graph exemplifying temporal change of magnetic signal strength in analyzing inter-molecular interaction (the horizontal axis expresses the time, the vertical axis expresses the magnetic signal strength).

[FIG. 3] FIG. 3 is an explanatory drawing showing a correlation diagram (reaction model) of the first molecule 1 (pKA), the second molecule 2 (quelcetin Q), and the third molecule 3 (HSA) in Example 1.

[FIG. 4] FIG. 4 is a graph showing temporal change of magnetic signal strength in Example 1 (the horizontal axis expresses the time, the vertical axis expresses the magnetic signal strength).

[FIG. 5] FIG. 5 is a graphic chart showing a reaction model and reaction rate expressions (Expressions (3) to (5)) of a competitive reaction in Example 1.

[FIG. 6] FIG. 6 is a graph showing temporal change of a magnetic signal obtained by a method in which the reaction is divided into two steps (the horizontal axis expresses the time, the vertical axis expresses the magnetic signal strength).

[FIG. 7] FIG. 7 is a graph showing an example of a practical strength range of a magnetic particle MP which can be used from a detection lower limit value and a detection upper limit value of a magnetic sensor MS and the magnetic particle MP (the horizontal axis expresses the time, the vertical axis expresses the magnetic signal strength).

[FIG. 8] FIG. 8 is a drawing showing an analysis result of inter-molecular interaction of the first molecule 1, the second molecule 2, and the third molecule 3 related to Example 2, and is a graph showing an example of temporal change of the magnetic signal strength (the horizontal axis expresses the time, the vertical axis expresses the magnetic signal strength).

[FIG. 9] FIG. 9 is a schematic drawing showing an embodiment of a device for analyzing inter-molecular interaction 100.

[FIG. 10] FIG. 10 is a schematic drawing explaining a block configuration and a flow of the device for analyzing inter-molecular interaction 100 related to another embodiment.

[FIG. 11] FIG. 11 is a flowchart showing a process flow of a program for analyzing inter-molecular interaction P along with a schematic configuration of the device for analyzing inter-molecular interaction 100.

[Description of Embodiments] [0010]

A method, a device, and a program for analyzing inter- molecular interaction related to an embodiment of the present invention will be hereinafter explained in detail referring to drawings from time to time. [0011] [An embodiment of a method for analyzing inter-molecular interaction]

In the drawings to be referred to, FIG. 1 is a graphic chart showing a flow of a method for analyzing inter-molecular interaction related to an embodiment (will be hereinafter referred to as "the present method") and contents in each step. That is to say, FIG. 1 shows an outline of an assay protocol that can evaluate inter-molecular interaction between a first molecule 1 fixed to a magnetic sensor MS and a second molecule 2 having a property of specifically bonding to the first molecule 1 and an influence rate of a third molecule stimulating inter-molecular interaction between the first molecule 1 and the second molecule 2. Although a competitive assay protocol where the third molecule 3 affects the second molecule 2 is shown in this FIG. 1, the present invention is not limited to it and can be applied also to a case where the third molecule 3 affects the first molecule 1.

[0012]

Also, the first step S1 shown in FIG. 1 schematically illustrates the magnetic sensor MS, the first molecule 1, and the first molecule 1 fixed to the magnetic sensor MS. The second step S2 shown in FIG. 1 schematically illustrates inter-molecular interaction between the first molecule 1 and the second molecule 2 having a property of specifically bonding to the first molecule 1 and an aspect that a first magnetic signal Sg1 is generated by that the magnetic sensor MS and a magnetic particle MP come close to each other sufficiently through the first molecule 1 and the second molecule 2 and temporal change of the first magnetic signal Sg1 is acquired. The third step S3 shown in FIG. 1 schematically illustrates an aspect that the third molecule 3 affecting the second molecule 2 as an example is added and temporal change of a second magnetic signal Sg2 having been generated is acquired. The fourth step S4 shown in FIG. 1 schematically illustrates that a tendency T1 of temporal change of the first magnetic signal Sg1 and a tendency T2 of temporal change of the second magnetic signal Sg2 having been acquired are compared with each other.

[0013] (From the first step S1 to the fourth step S4)

As shown in FIG. 1, the present method includes the first step S1 to the fourth step S4, and respective steps are executed in this order.

The first step S1 is a step for fixing the first molecule

1 to the surface of the magnetic sensor MS.

The second step S2 is a step for bonding the first molecule 1 and the second molecule 2 to each other and obtaining temporal change of the first magnetic signal Sg1, the second molecule 2 being labeled by the magnetic particle MP.

The third step S3 is a step for adding the third molecule 3 (a molecule other than the first molecule 1 and the second molecule 2 described above) at prescribed timing after starting bonding the first molecule 1 and the second molecule

2 to each other, and obtaining temporal change of the second magnetic signal Sg2.

[0014]

The fourth step S4 is a step for comparing the tendency T1 of temporal change of the first magnetic signal Sg1 and the tendency T2 of temporal change of the second magnetic signal Sg2 to each other. Also, in the fourth step S4, when these tendencies T1 and T2 of temporal change are different from each other, it is analyzed that the third molecule 3 affects inter-molecular interaction between the first molecule 1 and the second molecule 2. Also, in the fourth step S4, when these tendencies T1 and T2 of temporal change are the same with each other, it is analyzed that the third molecule 3 does not affect inter-molecular interaction between the first molecule 1 and the second molecule 2.

[0015]

As the tendencies T1 and T2, for example, a bonding constant, bonding rate constant, dissociation constant, dissociation rate constant, reaction rate parameter such as an initial reaction rate, and so on can be cited. Also, as the tendencies T1 and T2, for example, a temporal change curve obtained based on temporal change of a magnetic signal, its curvature or inclination, and the like can be cited. Further, as the tendencies T1 and T2, for example, inclination of a straight line in a case temporal change obtained based on temporal change of a magnetic signal is drawn by the straight line, and so on can be cited. Also, as the tendencies T1 and T2, for example, a reaction rate expression and the like can be cited. However, the tendencies T1 and T2 are not limited to them, and any one can be employed as far as it can be compared mutually with a same reference. Also, with respect to inclination of the temporal change curve, for example, by connecting two points of intersection obtained from the starting point and finishing point of a prescribed unit time such as 10 seconds or one minute after starting reaction and the temporal change curve, a straight line is obtained. In the present embodiment, inclination of the straight line thus obtained can be made "inclination of temporal change curve".

[0016]

(Manufacturing of the magnetic sensor device MSD and the like)

According to the present embodiment, one where the first molecule 1 is fixed to the surface of the magnetic sensor MS is a magnetic sensor device MSD. Here, "to be fixed" means to be bonded stably. Also, "to be bonded stably" means that the first molecule 1 and the surface of the magnetic sensor MS maintain a spatial position with each other under a use condition namely under an assay condition for example for a period longer than a transient period. Therefore, the first molecule 1 and the surface of the magnetic sensor MS can be bonded to each other stably by non-covalent bonding or by covalent bonding. As examples of non-covalent bonding, non- specific adsorption, bonding based on static electricity (for example, ion interaction, ionic pair interaction), hydrophobic interaction, hydrogen bonding interaction, specific bonding by a specific first molecule 1 bonded to the support surface by covalent bonding, and so on can be cited. As an example of covalent bonding, covalent bonding formed between a functional group that is present on the surface of the magnetic sensor MS namely a hydroxyl group (-OH group) for example and the first molecule 1 can be cited. The functional group may be one formed naturally, or may be present as an element of the bonding group having been introduced. Therefore, the first molecule 1 may be adsorbed to the surface of the magnetic sensor MS, may be adsorbed physically, may be adsorbed chemically, or may be bonded to the surface of the magnetic sensor MS by covalent bonding. [0017]

As shown in the first step S1 of FIG. 1, the present method includes manufacturing of the magnetic sensor device MSD where the first molecule 1 is fixed to the surface of the magnetic sensor MS. Also, as shown in the second step S2 of FIG. 1, the first molecule 1 of the magnetic sensor device MSD having been manufactured and the second molecule 2 labeled by the magnetic particle MP bond to each other. The first molecule 1 and the second molecule 2 are element molecules (bonding pair elements) of inter-molecular interaction which become analysis objects. The magnetic sensor device MSD generates an electric signal (magnetic signal) according to the magnetic particle MP that comes sufficiently close to the magnetic sensor MS through bonding the first molecule 1 and the second molecule 2 with each other. Also, the magnetic particle MP may be a portion or a region within the second molecule 2, or may be a separate molecule (particle) bonded to the second molecule 2. When the magnetic particle MP is a particle separate from the second molecule 2, the magnetic particle MP can bond stably to the second molecule 2 by covalent bonding or by non-covalent bonding for example. [0018]

In manufacturing the magnetic sensor device MSD, the first step SI may include various different appropriate process sub- combinations from a viewpoint when the first molecule 1 and the second molecule 2 bond to each other, or from a viewpoint of a configuration of the first molecule 1 with respect to the magnetic sensor MS, a configuration of the second molecule 2 with respect to the magnetic sensor device MSD, and so on for example. Here, as an appropriate process sub-combination, for example, to allow different capturing probe or molecule bonded to the magnetic sensor MS to be bonded so that the magnetic sensor MS is bonded specifically to a separate molecule, and so on can be cited.

[0019]

(Analysis objects of inter-molecular interaction: the first molecule 1, the second molecule 2, the third molecule 3) Inter-molecular interaction analyzed by the present method may be configured of molecules of a bonding pair which exert selectively or specifically high affinity with each other. Molecules (the first molecule 1 and the second molecule 2) of the bonding pair are biological molecules of the kind different from each other such as a ligand and a receptor for example, and both of them may be biological macromolecules, and may not be biological macromolecules.

[0020]

The term "ligand" used in the present description means a portion/molecule that can bond a molecule becoming a bonding pair by covalent bonding or in a chemical manner. The ligand may be naturally derived one, or may be synthesized one. The ligand can allow the magnetic sensor MS or the magnetic particle MP which are a bonding object to bond thereto directly or through prescribed bonding matter by covalent bonding or by non-covalent bonding.

As the ligand, for example, a medical agent, an agonist and antagonist for a cell membrane receptor, toxin and venom, viral epitope, hormone, opiate, steroid, peptide, enzyme substrate, capture agent, lectin, sugar group, oligonucleotide, nucleic acid, oligosaccharide group, protein, polyphenol group, and the like can be cited, however, the ligand is not limited to them. [0021]

The term "receptor" used in the present description means a portion/molecule having affinity with the ligand. The receptor may be naturally derived one, or may be synthesized one. Also, the receptor may be used in a native state, and may be used in a modified state such as an aggregate with other species. Either one of the first molecule 1 and the second molecule 2 may be a ligand or may be a receptor. That is to say, when one is a ligand, the other becomes a receptor. The receptor can allow the magnetic sensor MS or the magnetic particle MP which are the bonding objects to bond directly or through prescribed bonding matter thereto by covalent bonding or by non-covalent bonding. As the bonding matter, for example, polyethyleneglycol (PEG), peptide linker, DNA linker, and the like can be cited. As the receptor, for example, a medical agent, antibody, cell membrane receptor, monoclonal antibody and antiserum having a prescribed antigenic determinant group, virus, cell, polynucleotide, nucleic acid, peptide, capture agent, lectin, sugar group, polysaccharide group, cell membrane, cell organ, and the like can be cited, however, the receptor is not limited to them. The receptor may be called an anti-ligand in a technical field to which the present invention belongs.

[0022]

In a certain case, the molecule of a bonding pair may be substantially different according to inter-molecular interaction of the analysis object. The inter-molecular interaction described above occurs by specificity between molecules of a bonding pair under an environmental condition of interaction. The inter-molecular interaction of the analysis object also includes any interaction between molecules of a bonding pair. As an example of such inter- molecular interaction, for example, nucleic acid hybridization interaction, inter-protein interaction, protein-nucleic acid interaction, enzyme-substrate interaction, receptor-ligand interaction, antibody-antigen interaction, receptor-agonist or antagonist interaction, and so on can be cited, however, such inter-molecular interaction is not limited to them. At least one or any one selected from them can be made an object of the inter-molecular interaction. [0023]

Also, as an example of a molecule having the inter- molecular interaction described above, there are a biological macromolecule and a small molecule, and the small molecule may be an organic small molecule or an inorganic small molecule. "Biological macromolecule" may be a macromolecule with one or plural types of repeating unit, or may be one without a repeating unit. The biological macromolecule may be formed synthetically, or may be biologically derived. The biological macromolecule only has to be configured of or contain amino acid analog, non-amino acid group, nucleotide analog, or non- nucleotide group in addition to peptide, polynucleotide, polysaccharide, and the like.

[0024]

Also, the present embodiment is characterized in that influence of the third molecule 3 on the bonding pair of the first molecule 1 and the second molecule 2 can be analyzed. Therefore, all chemical matter and biological matter affecting various kinds of inter-molecular interaction exemplified above become an object of the third molecule 3. Also, the third molecule 3 is not limited to one formed of one component, and may be one where plural components are mixed. As the third molecule 3, for example, a molecule medical agent, an agonist and antagonist for a cell membrane receptor, toxin and venom, viral epitope, hormone, opiate, steroid, peptide, enzyme substrate, capture agent, lectin, sugar group, oligonucleotide, nucleic acid, oligosaccharide group, protein, polyphenol group, and the like can be cited, however, the third molecule 3 is not limited to them.

[0025]

(Magnetic sensor device MSD)

As described above, the magnetic sensor device MSD generates an electric signal (magnetic signal) according to the magnetic particle MP that comes sufficiently close to the magnetic sensor MS through the first molecule 1 and the second molecule 2. It is preferable that the magnetic sensor MS is one measuring the change amount of the magnetic resistance. Therefore, as the magnetic sensor MS, for example, known giant magnetic resistance (GMR) sensor can be used, however, the magnetic sensor MS is not limited to it. For example, as the magnetic sensor MS, known spin valve detector, or a magnetic tunnel joint (MTJ) detector (TMR sensor) can also be used. When such a magnetic sensor MS is used, the magnetic signal is surely obtained.

[0026]

A magnetic field is applied to the magnetic sensor MS and a solution containing the magnetic particle MP that is sufficiently close to the magnetic sensor MS by an external magnetic field generation device (not illustrated), the magnetic particle MP having superparamagnetism is thereby magnetized, and the strength of the induced magnetic field can be detected by the magnetic sensor MS. As the external magnetic field generation device, for example, a Helmholtz coil can be used. The Helmholtz coil is configured that identical two coils are disposed to have a same axis. Since the strength of a magnetic field attenuates with the cube of the distance, only the magnetic particle MP that is present within approximately 500 nm from the surface of the magnetic sensor MS and is fixed for a time sufficient for measurement is detected. When a GMR sensor is used for the magnetic sensor MS, the resistance value of the magnetic sensor MS is measured by a measuring instrument (not illustrated), the measured value is taken to a controller (not illustrated), and calculation, storage of data, display, and the like are executed .

[0027]

The magnetic sensor device MSD can have various different configurations irrespective of whether an analysis device used employs either of a batch type and a flow through type. Therefore, the magnetic sensor device MSD can have a freely- selected configuration as far as the magnetic sensor MS being sufficiently close to the magnetic particle MP through the bonding pair elements of inter-molecular interaction (the first molecule 1 and the second molecule 2) and generating a magnetic signal in accordance with the magnetic particle MP is provided. Therefore, for example, the magnetic sensor device MSD can have a well construction where the magnetic sensor MS is arranged in the bottom part or the wall part of a fluid storage structure such as a microtiter plate. Also, for example, the magnetic sensor device MSD can have a flow through construction where the magnetic sensor MS is arranged in the wall part of a flow cell that involves flowing in and flowing out of fluid.

[0028]

In an embodiment, the magnetic sensor device MSD may include two or more separate magnetic sensors MS on the surface of the substrate. Also, in an embodiment, the magnetic sensor device MSD may include an array of the magnetic sensor MS on the surface of the substrate.

[0029]

"Array" includes a freely-selected two-dimensional array with an addressable area namely a spatially addressable area for example. When plural number of the magnetic sensors MS are disposed at prescribed positions (namely "address") on the array, the array is "addressable". The features of the array (namely plural number of the magnetic sensors MS) may be arranged to be apart from each other by a space interposed therebetween .

[0030]

The substrate described above may support 1, 2, 4, or more arrays. Any one or all of arrays that can detect a same or a different analysis object may include plural number of separate magnetic sensors MS. The array may include the magnetic sensors MS of 2 or more, 4 or more, 8 or more, 10 or more, 50 or more, 100 or more, 1,000 or more, 10,000 or more, or 100,000 or more. In an embodiment, it is preferable that the array includes 2 or more plural number of separate magnetic sensors MS. For example, 16 pieces of the magnetic sensors MS can be disposed in each of the area of the array that includes 2x2 addressable areas.

[0031]

In an embodiment, the magnetic sensor MS can be disposed in an array having 10 cm ² or less, 5 cm ² or less, 1 cm ² or less, 50 mm ² or less, 20 mm ² or less, 10 mm ² or less, or further smaller area for example. Also, the magnetic sensor MS can have a size of the range of 10 μmx10 μm to 200 μmx200 μm for example, and can more preferably have the size of 100 μmx100 μm or less, 90 μmx90 μm or less, and 50 μmx50 μm or less.

[0032]

In an embodiment, the first molecule 1 can be changed according to the nature of inter-molecular interaction analysis to be executed. For example, although only the first molecule 1 is solid-phased, it becomes important that the recognition site of inter-molecular interaction=epitope position and 3D structure are not affected even when the first molecule 1 is solid-phased. Therefore, out of three molecules that can be set as the first molecule 1, the second molecule 2, and the third molecule 3, which should be made to be the first molecule 1 can be set according to the property of each molecule .

The first molecule 1 may be a molecule of inter-molecular interaction of the analysis object or a molecule getting involved in inter-molecular interaction of the analysis object. That is to say, the first molecule 1 may be a capturing probe and the like that is bonded to the second molecule 2 to which the magnetic particle MP is bonded. [0033]

When the magnetic sensor device MSD includes two sets or more magnetic sensors MS, each magnetic sensor MS may include an identical or different first molecule 1 that is bonded to the surface of each magnetic sensor MS. By doing so, as the Multiplex analysis, interaction between different molecules can be analyzed. Therefore, in order that each magnetic sensor MS is specifically bonded to a separate molecule, a different capturing probe or molecule bonded to the magnetic sensor MS may be present on the sensor surface of such a device. The magnetic sensor device MSD may further include the magnetic sensor MS to which the first molecule 1 is not bonded. For example, such a magnetic sensor MS to which the first molecule 1 is not bonded can function as a source of a reference electric signal or a control electric signal. That is to say, since the first molecule 1 is not bonded thereto, such a magnetic sensor MS can be used in quality control of watching a non-singular signal of magnetic sensor device MSD + magnetic particle MP. [0034]

In the magnetic sensor device MSD including plural number of the magnetic sensors MS, a space area between the magnetic sensors MS where the magnetic sensor MS and the first molecule 1 are not bonded to each other may be present. Such a space area may have various sizes and configurations. In some cases, such a space area may be configured to reduce or prevent fluid movement between different magnetic sensors MS. Such space area may be configured of a freely-selected hydrophobic material and/or a fluid barrier (like a wall) for example. [0035]

With respect to the basic method and mechanism for reading a magnetic signal from the magnetic sensor MS, it is possible to apply such a general one as described in already reported literatures (for example, "Magneto-nanosensor platform for probing low-affinity protein-protein interactions and identification of a low-affinity PD-L1/PD-L2 interaction", Nature Commun., 7, 12220, 2016).

[0036]

(Magnetic particle MP)

In the present embodiment, a freely-selected magnetic particle MP can be used. When the magnetic particle MP is sufficiently related (brought close) to the magnetic sensor MS through the first molecule 1 and the second molecule 2, the magnetic particle MP is detected by the magnetic sensor MS and allows the magnetic sensor MS to output an electric signal. When the distance between the center of the magnetic particle

MP and the surface of the magnetic sensor MS is 200 nm or less, preferably 100 nm or less, and more preferably 50 nm or less, the magnetic particle MP can get close sufficiently to the magnetic sensor MS, and can generate a magnetic signal. [0037]

In an embodiment, the magnetic particle MP is a nanoparticle. A nanoparticle useful in implementing an embodiment is of a magnetic (ferromagnetic for example) colloidal material and particle (magnetic nanoparticle). The magnetic particle MP can be made a magnetic nanoparticle of high momentum, and may be a superparamagnetic or synthesized antiferromagnetic nanoparticle including two or more layers of a ferromagnetic body of high momentum bonded in an antiferromagnetic manner. All the nanoparticles of these types are less likely to be agglomerated since they are "non- magnetic" in a state where a magnetic field is not present. [0038]

Also, in an embodiment, the magnetic particle MP may have slight residual magnetism if it is of a degree of not agglomerated within a solution. As an example of the magnetic particle MP having slight residual magnetism, there is a superparamagnetic particle and an antiferromagnetic particle. In some cases, the magnetic particle MP has magnetic moment detectable under a magnetic field of approximately 100 Oe. Also, in some cases, the size of the magnetic particle MP is made the same as or less than the size of a biological molecule of the analysis object. By doing so, since the magnetic particle MP is not too large, it is considered that the magnetic particle MP is less likely to interfere with inter-molecular interaction of the analysis object. In some cases, the magnetic particle MP has a generally same shape so as to be easily used during analysis, and is chemically stable under a biological environment. In some cases, the magnetic particle MP has biological adaptability (namely water- soluble), and is functionalized so as to be easily bonded to a receptor that specifically bonds to a biological molecule of an object namely an object specimen for example. [0039]

In an embodiment, the size of the magnetic particle MP for example is 5 nm to 300 nm in diameter, preferably 5 nm to 250 nm in diameter, more preferably 5 nm to 150 nm in diameter, and further more preferably 5 nm to 20 nm in diameter. The particle diameter can be confirmed by a Zetasizer. With respect to the size of the magnetic particle MP, for example, a magnetic particle MP whose diameter is 5 nm, 10 nm, 15 nm, 20 nm, 25 nm, 50 nm, 80 nm, 100 nm, 150 nm, 250 nm, or 300 nm and a magnetic particle MP whose diameter falls within a range of freely-selected two values out of these values are suitable to the use in the present method. Also, when the diameter of the magnetic particle MP is smaller than 5 nm, since the magnetic signal strength is weak, the sensitivity may deteriorate. On the other hand, when the diameter of the magnetic particle MP becomes larger than 300 nm, since the mass per a particle becomes too large, the magnetic signal strength may extremely deteriorate. Further, although the shape of the magnetic particle MP suitable to the use in the present embodiment is spherical, the shapes of a disk, a bar, a coil, a fiber, and the like are also possible.

[0040]

In an embodiment, the second molecule 2 and the magnetic particle MP are stably bonded to each other. "Stably bonded" means that the second molecule 2 and the magnetic particle MP maintain a spatial position with each other for a period longer than a transient period under a use condition namely an assay condition for example. Therefore, the second molecule 2 and the magnetic particle MP can be bonded to each other stably by non-covalent bonding or by covalent bonding. As examples of non-covalent bonding, there are non-specific adsorption, bonding based on static electricity (for example, ion interaction, ionic pair interaction), hydrophobic interaction, hydrogen bonding interaction, specific bonding by a specific second molecule 2 bonded to the support surface by covalent bonding, and so on. As an example of covalent bonding, peptide bonding can be cited. Peptide bonding can be implemented, for example, utilizing a cross-linking agent such as 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) when there is an amino group (NH ₂ group) on the second molecule 2 with respect to a functional group namely carboxylate ester (COO group) for example present on the surface of the magnetic particle MP. By doing so, the magnetic particle MP can be introduced to the second molecule 2 by a strong and highly versatile means. The functional group may be one formed naturally, or may be present as an element of the bonding group having been introduced as described above. [0041]

Also, when the second molecule 2 is to be labeled by the magnetic particle MP, it is preferable that magnetization per one particle becomes even. When the second molecule 2 is to be labeled by the magnetic particle MP, known and freely-selected labeling technology can be applied if stability and functionality of the second molecule 2 and the magnetic particle MP can be maintained. Also, considering that the magnetic particle MP is labeled to the second molecule 2 which is the analyte, it is preferable to use the magnetic particle MP where a reactive group is evenly introduced to the surface. [0042] (Acquisition of an electric signal (magnetic signal) from the magnetic sensor MS)

As described above, according to the present embodiment, an electric signal namely a magnetic signal can be obtained according to the magnetic particle MP that comes sufficiently close to the magnetic sensor MS through bonding the first molecule 1 and the second molecule 2 to each other. In an embodiment, this magnetic signal is a real-time signal, and is configured of two or more data points obtained over a prescribed time. For example, this magnetic signal is configured of a set of continuous data points (in a trace form for example) obtained continuously over a prescribed time. The prescribed time can be selected between 1 second to 10 hours, preferably between 10 seconds to 1 hour, and more preferably between 1 minute to 15 minutes for example. The number of the data points of the real-time signal can be changed optionally. Also, in some cases, the number of the data points is not limited specifically as far as it is sufficient to produce the data of a continuous range over a time course of the real-time signal.

[0043] (Quantitative determination of reaction rate parameter from real-time signal)

According to the present embodiment, the tendency of temporal change of the magnetic signal is obtained as described above. As an example, according to the present embodiment, following acquisition of the real-time signal, the reaction rate parameter (bonding constant, dissociation constant, bonding rate constant, dissociation rate constant, and the like for example) of the inter-molecular interaction is determined quantitatively from the real-time signal. In other words, the real-time signal is used for quantitatively determining the reaction rate parameter of the analysis obj ect so that the reaction rate parameter of the analysis obj ect can be obtained from the real-time signal .

[ 0044 ]

In some cases , the reaction rate parameter of the analysis obj ect is determined quantitatively by that the real-time signal is processed using an adaptive algorithm . The adaptive algorithm means a set of rules for determining the reaction rate parameter of an obj ect by applying an expression to one or plural number of the real-time signal obtained from a prescribed assay for example . The adaptive algorithm only has to be capable of determining the reaction rate parameter of the analysis obj ect , and any form can be used .

[ 0045 ]

( Test specimen)

With respect to the test specimen applied to the present method, there is no speci fic limitation as far as it is a specimen possibly containing material to be tested which becomes the measurement obj ect . As the test specimen, for example , specimens of the biological origin such as blood, blood plasma, blood serum, cerebral spinal fluid, amnion fluid, milk, sweat , urine , saliva, sputum, stercus , tissue , cell culture of the animal origin can be cited . Further, although these test specimens can be made the measurement obj ects as they are , they may be subj ected to pretreatment such as separation, extraction, refining of the material to be tested .

[ 0046 ]

(More detailed explanation on the present method)

The present method will be hereinafter explained more concretely and in detail further referring to FIG . 1 .

As shown in FIG . 1 , in the first step S1, the first molecule 1 is fixed onto the surface of the magnetic sensor MS . Such fixing can be executed by any means as far as chemical matter and biological matter can be retained to the magnetic sensor MS for a constant time such as a covalent boding method, hydrophobic bonding method, ion bonding method, biotin-avidin interaction, and antigen-antibody reaction . Such fixing is executed by a user . [ 0047 ]

Prior to the second step S2 , the second molecule 2 is labeled by the magnetic particle MP . Labeling by the magnetic particle MP is executed by a user using a labeling reagent that is generally made into a kit . [ 0048 ]

Next , in the second step S2 , the magnetic sensor MS described above to which the first molecule 1 is fixed and the second molecule 2 described above to which the magnetic particle MP is labeled are made to be in contact with each other . Thus , inter-molecular interaction 12 occurs between the first molecule 1 and the second molecule 2 , and a complex such as an immune complex for example is formed . The method for allowing them to be in contact with each other may be a general assay method such as a batch type using a microtiter plate and the like and a flow type using a micro flow passage and the like . In the batch type , it is preferable that the second molecules 2 are dispersed with such capacity that the second molecules 2 that are constantly fresh are circulated with respect to the first molecules 1 that are constantly fixed to the magnetic sensors MS and evenly . In the flow type of a micro flow passage and the like , it is preferable that , in allowing fluid to flow, the flow rate is made to adapt to the volume of the micro flow passage so that a laminar flow is not generated . In either of the batch type and the flow type , when these preferable aspects are employed, reaction rate expressions described below can be applied suitably to inter- molecular interaction between the first molecule 1 and the second molecule 2 .

[ 0049 ]

As shown in the second step S2 , when the first molecule 1 and the second molecule 2 are brought into contact with each other and a complex is formed, the magnetic particle MP comes close to the magnetic sensor MS resultantly, and therefore the magnetic signals detected on the magnetic sensor MS increase according to the volume of the complex . [ 0050 ]

In the third step S3 , after a constant time ( at prescribed timing) after the first molecule 1 and the second molecule 2 are made to be in contact with each other, the third molecule 3 is added . The method for adding the third molecule 3 may be a general method such as a micropipette and a dispenser .

Also , with respect to the timing for adding the third molecule 3 , in order that the inter-molecular interaction between the first molecule 1 and the second molecule 2 can be analyzed highly precisely, it is preferable that the magnetic signal strength obtained from them is high to some degree . Further, the timing is preferable to be within such a range that the magnetic signal by bonding the second molecule 2 and the third molecule 3 to each other does not saturate after adding the third molecule 3 . Therefore , in determining the timing, it is preferable to grasp beforehand the fixing amount of the first molecule 1 , the density of the second molecule 2 , the labeling ef ficiency of the magnetic particle MP to the second molecule 2 , and the magnetic signal strength by bonding the second molecule 2 and the third molecule 3 to each other . [ 0051 ]

Here , it is preferable to execute the second step S2 and the third step S3 as follows .

That is to say, in the second step S2 , it is preferable that the magnetic signal of the magnetic particle MP labeled to the second molecule 2 is acquired on real-time from the magnetic sensor MS as the first magnetic signal Sg1 , and that the tendency T1 of temporal change of the first magnetic signal Sg1 is acquired . Also , in the third step S3 , it is preferable that the magnetic signal of the magnetic particle MP labeled to the second molecule 2 is acquired on real-time from the magnetic sensor MS as the second magnetic signal Sg2 , and that the tendency T2 of temporal change of the second magnetic signal Sg2 is acquired .

When the second step S2 and the third step S3 are thus executed, the inter-molecular interaction can be analyzed quantitatively and dynamically . [ 0052 ]

Also , in the fourth step S4 to be executed next , the tendency T1 before adding the third molecule 3 and the tendency T2 after adding the third molecule 3 are compared with each other . Therefore , in the third step S3 , it is preferable to add the third molecule 3 so that the di f ference of the final density of the second molecule 2 between before and after adding the third molecule 3 becomes a volume that can allow the influence . For example , when the adding volume of the third molecule 3 is 1 / 100 of the volume of the second molecule 2 , drop of the final density of the second molecule 2 remains 1 % , and therefore the influence on the di f ference of the final density can be allowed . [ 0053 ]

When the third molecule 3 reacts with the second molecule 2 to be bonded, and so on in the third step S3 , inter- molecular interaction 13 occurs between the third molecule 3 and the second molecule 2 . By the influence , the inter- molecular interaction 12 having occurred in the second step S2 changes to inter-molecular interaction 12a . When the third molecule 3 exhibits a competitive nature , the tendency T2 of temporal change of the second magnetic signal Sg2 of the inter-molecular interaction 12a (bonding constant , dissociation constant , bonding rate constant , dissociation rate constant , and the like for example ) becomes smaller than the tendency T1 of temporal change of the first magnetic signal Sg1 of the inter-molecular interaction 12 . On the other hand, when the third molecule 3 exhibits a promotional nature , the tendency T2 of temporal change of the second magnetic signal Sg2 of the inter-molecular interaction 12a (bonding constant , dissociation constant , bonding rate constant , dissociation rate constant , and the like for example ) becomes larger than the tendency T1 of temporal change of the first magnetic signal Sg1 of the inter-molecular interaction 12 .

[ 0054 ]

Here , FIG . 2 is a graph exempli fying temporal change of the magnetic signal strength in analyzing inter-molecular interaction . FIG . 2 shows in a simpli fied manner temporal change of the magnetic signal obtained with respect to a case the third molecule 3 has a competitive nature and a promotional nature and a case the third molecule 3 is not added .

As described above , according to the present method, the second step S2 is started in analyzing inter-molecular interaction, and temporal change of the first magnetic signal Sg1 is obtained. As described above, in an embodiment, since the magnetic signal (the first magnetic signal Sg1) is a real- time signal, as an example of the tendency T1, a first temporal change curve C1 for example as shown in FIG. 2 is obtained .

Also, the third molecule 3 is added at prescribed timing 23 to start the third step S3, and a temporal change of the second magnetic signal Sg2 is obtained. Similarly to the above, in an embodiment, since the magnetic signal (the second magnetic signal Sg2) is a real-time signal, as an example of the tendency T2, a second temporal change curve C2 for example as shown in FIG. 2 is obtained.

[0055]

At this time, when the third molecule 3 has a competitive nature, the tendency T2 (namely a second temporal change curve C2c and inclination, bonding constant, bonding rate constant, and the like) after adding the third molecule 3 becomes smaller than the tendency of temporal change (namely a second temporal change curve C2n and inclination, bonding constant, bonding rate constant, and the like) of a case of not adding the third molecule 3.

In contrast, when the third molecule 3 has a promotional nature, the tendency T2 (namely a second temporal change curve C2a and inclination, bonding constant, bonding rate constant, and the like ) after adding the third molecule 3 becomes larger than the tendency of temporal change (namely the second temporal change curve C2n and inclination, bonding constant , bonding rate constant , and the like ) of a case of not adding the third molecule 3 .

Also , when the third molecule 3 does not af fect the inter- molecular interaction 12 between the first molecule 1 and the second molecule 2 , the tendency T2 (namely the second temporal change curve C2n and inclination, bonding constant , bonding rate constant , and the like ) after adding the third molecule 3 becomes similar to the tendency of temporal change of a case of not adding the third molecule 3 . [ 0056 ]

Also , it is preferable intrinsically that the inter- molecular interaction ( tendency) of a case of not adding the third molecule 3 is the same as the inter-molecular interaction 12 ( the tendency T1 ) before adding the third molecule 3 . That is to say, it is preferable that they are on the bases of the same reaction rate expression, bonding constant , and bonding rate constant , and it is also preferable that the temporal change curves ( the first temporal change curve C1 , the second temporal change curve C2n) obtained become the same with each other .

Also , when data on the inter-molecular interaction 12 ( the first temporal change curve C1 , reaction rate expression, bonding constant , bonding rate constant , and the like for example) and data on the inter-molecular interaction 12a (the second temporal change curves C2a, C2c, reaction rate expression, bonding constant, bonding rate constant, and the like for example) are confirmed respectively beforehand, it is preferable since reliability can be confirmed in executing the present method.

[0057]

Explanation will be continued returning to FIG. 1. As shown in FIG. 1, in the fourth step S4, the tendency T1 of temporal change of the fist magnetic signal Sg1 and the tendency T2 of temporal change of the second magnetic signal Sg2 are compared to each other (T1=T2?) . Also, as described above, in the fourth step S4, when these tendencies T1, T2 of temporal change are different from each other, it is analyzed that the third molecule 3 affects the inter-molecular interaction 12 between the first molecule 1 and the second molecule 2. Further, in the fourth step S4, when these tendencies T1, T2 of temporal change are the same with each other, it is analyzed that the third molecule 3 does not affect the inter-molecular interaction 12 described above. [0058] (Acquisition of inter-molecular interaction (rate constant) from temporal change of magnetic signal)

From the first temporal change curve C1 and the second temporal change curve C2, the rate constant (bonding rate constant, dissociation rate constant) which is one parameter of analysis of inter-molecular interaction can be calculated.

As a method for calculating the rate constant, the Langmuir adsorption expression for example can be applied. In applying the Langmuir adsorption expression, the rate constant can be calculated from coefficients S ₀ and a extracted by expressions (1) , (2) below by global fitting. That is to say, the bonding process and the dissociation process of the first molecule 1 and the second molecule 2 are in accordance with the expressions (1) , (2) below. [0059] (Bonding process)

S t=S ₀ x { 1-exp ( -k _on ■ C ₀ ■ t ) } (1)

Here, in the expression (1) , St is the bonding strength, So is the saturated density (namely the saturated density when the first molecule 1 and the second molecule 2 bond to each other) detectable on the magnetic sensor MS, k _on is the bonding rate constant, Co is the initial density (mol/L) of the second molecule 2, and t is the temperature (°C or K) . By solving the expression (1) with respect to k _on , the bonding rate constant can be obtained. [0060] (Dissociation process)

St=a ■exp (-k _off ■ t) (2)

Here, in the expression (2) , St is the bonding strength, a is the magnetic signal at the time of starting the dissociation reaction, k _off is the dissociation rate constant, and t is the temperature (°C or K) . By solving the expression

(2) with respect to k _off , the dissociation rate constant can be obtained .

[0061]

Example 1 related to an embodiment of the present method described above will be shown below. Also, the contents of the present method are not limited by any means by this Example 1. [0062] (Example 1) [I] In Example 1, as the magnetic sensor MS, a conventional Giant Magneto-Resistance Sensor (will be hereinafter expressed as a GMR sensor or expressed simply as GMR) was used. As the first molecule 1, cAMP-dependent Protein Kinase (will be hereinafter referred to as pKA) was used. As the second molecule 2, quercetin labeled by the magnetic particle MP was used. As the third molecule 3, Human Serum Albumin (will be hereinafter referred to as HSA) was used. [0063]

That is to say, Example 1 shows an example of analyzing interaction of the first molecule 1 (pKA) , the second molecule 2 (quercetin) , and the third molecule 3 (HAS) . The interaction is caused by that the third molecule 3 (HAS) initiates an inhibition reaction with the second molecule 2 (quercetin) . [0064] (1) As described below, pKA (made by BioLabs Inc.) which is the first molecule 1 is fixed onto the surface of the magnetic sensor MS.

First, pKA was molten in Phosphate Buffer Saline (will be hereinafter referred to as PBS) of pH 7.4 so that the final density of pKA became 50 pg/mL. Also, the solution was coated by 1.5 nL on a GMR sensor made by MagArray, Inc. Since the liquid volume was small, pKA was fixed by drying of several minutes or less. Thus, a pKA-fixed magnetic sensor (the magnetic sensor device MSD) was obtained in which pKa that was the first molecule 1 was fixed to the magnetic sensor MS. [0065] (2) As described below, a magnetic nanoparticle that was the magnetic particle MP was labeled to quercetin that was the second molecule 2.

First, quercetin was molten in 25%-dimethylsulfoxide/PBS to have the final density of 1 mg/mL, and a quercetin solution was obtained. 200 μL of the magnetic nanoparticles (Basic MicroBeads, Order No. 130-048-001) made by Miltenyi Biotec B.V. and 50 μL of the quercetin solution were mixed with each other, were stirred at 500 rpm for whole day and night, and a mixed solution was obtained. Thereafter, this mixed solution was applied to a separating refining column (p Column) made by Miltenyi Biotec B.V., was cleaned and extracted by PBS, thereby non-labeled quercetin was removed, and magnetic nanoparticle-labeled quercetin was obtained. [0066]

(3) Next, Lyphocheck Tumor Marker Plus Control, Level 3 made by BioRad Laboratories, Inc. was molten by 2 mL of the Distilled Water, and thereby 100% of an HAS solution containing HAS that was the third molecule 3 was obtained. [0067]

(4) The pKA-fixed magnetic sensor having been manufactured was attached to a magnetic signal measurement apparatus MR-813 made by MagArray, Inc., and was blocked for 15 minutes by 0.05% SDS . Thereafter, the magnetic nanoparticle-labeled quercetin was made to react with different HSA density conditions (0%, 0.01%, 0.1%) appropriately using the HSA solution. That is to say, the first molecule 1 (pKA) , the second molecule 2 (quercetin Q) , and the third molecule 3 (HAS) were made to collectively react with each other at the same time by one process.

[0068] FIG. 3, FIG. 4

A correlation diagram (reaction model) of the first molecule 1 (pKA) , the second molecule 2 (quercetin Q) , and the third molecule 3 (HAS) in this reaction is shown in FIG. 3. Also, temporal change of the magnetic signal strength is shown in FIG. 4. Further, the bonding rate constant k _on obtained by the global fitting method based on the magnetic signal strength of 20 minutes after starting of the reaction ("START" of FIG. 4) and this curve is shown in Table 1. [0069]

[Table 1]

[0070]

In general, when a competitive reaction occurs, since the density of a corresponding molecule actually related to bonding lowers, the magnetic signal strength should drop while the bonding rate constant k _on remains unchanged. However, as shown in Table 1, based on the reaction model shown in FIG. 3, although the magnetic signal strength dropped, the bonding rate constant k _on became high, and the calculation result turned out to be insufficiently accurate. This was considered to be attributable to that the first molecule 1 (pKA) , the second molecule 2 (quercetin Q) , and the third molecule 3 (HSA) were made to collectively react with each other at the same time by one process. [0071] FIG. 5 [II] Therefore, the present inventors assumed the reaction model of the competitive reaction and the reaction rate expressions (expressions (3) to (5) ) shown in the graphic chart of FIG. 5.

Based on these assumptions, the present inventors decided to add the third molecule 3 after a prescribed time elapsed from starting of the reaction of the first molecule 1 and the second molecule 2 in order to analyze the influence of HSA that was the third molecule 3 recognizably and precisely. That is to say, it was decided that the reaction was divided into such two steps that the first molecule 1 and the second molecule 2 were made to react with each other first, and the third molecule 3 was added thereafter.

[0072]

First, similarly to the method described in [I] , the pKA- fixed magnetic sensor (the magnetic sensor device MSD) was subjected to immerse treatment in a well containing the blocking solution (0.05% SDS) . Next, the pKA-fixed magnetic sensor was made to react in a well containing 400 μL of the magnetic nanoparticle-labeled quercetin including quercetin Q (the second molecule 2) . [0073] FIG. 6

Next, 20 minutes thereafter, the HSA solution containing HSA (the third molecule 3) was added with the same condition as that of the previous experiment (however, the HSA density condition was different: 0%, 0.01%, 0.02%, 0.04%) , and the magnetic signal was measured on real-time. Also, it was arranged so that the total density was not affected by adding the HSA solution. In concrete terms, the adding amount of the HSA solution was made 4 μL (=volume of 1/100 of solution of the magnetic nanoparticle-labeled quercetin) . Further, the present reaction was arranged so that quercetin (the second molecule 2) and HSA (the third molecule 3) were sufficiently stirred by oscillating the well. The result is shown in FIG. 6. FIG. 6 is a graph showing temporal change of a magnetic signal obtained by a method in which the reaction is divided into two steps.

Also, when the parameters (HSA initial density Pp _max , bonding rate constant k _on , dissociation rate constant k _b , bonding rate constant k _d ) obtained from the reaction curve shown in FIG. 6 were calculated based on the reaction rate expressions (expressions (3) to (5) ) shown in FIG. 5, the result was as shown in Table 2.

[0074]

[Table 2]

[0075]

(Result of Example 1)

As shown in Table 2, the initial density (Pp _max ) of HSA that was the third molecule 3 became larger in proportion to increase of the HSA density. Also, the magnetic signal strength dropped while the bonding rate constant k _on remained unchanged. This is according to the theory, and the calculation is considered to have been executed correctly .

[ 0076 ]

That means , as shown in Table 2 , after adding HSA, the magnetic signal strength dropped in proportion to the density of HSA. That is to say, it can be said that only the density of HSA ( the third molecule 3 ) af fected inter-molecular interaction between pKA ( the first molecule 1 ) and quercetin ( the second molecule 2 ) . In other words , it can be said that inter-molecular interaction between quercetin ( the second molecule 2 ) and HSA ( the third molecule 3 ) also could be analyzed in accordance with the reaction model and the rate expressions .

From the above , it was confirmed that , according to the present method, not only inter-molecular interaction between the first molecule 1 and the second molecule 2 which is general as the inter-molecular interaction but also the influence the third molecule 3 exerts to the inter-molecular interaction between these two molecules can be analyzed . [ 0077 ] (Application of Example 1 )

It is considered that , when the present method is applied, inter-molecular interaction with further multiple steps of not only adding elements of two stages of the second molecule 2 and the third molecule 3 shown in Example 1 but also adding a fourth molecule , a fi fth molecule , and the like can also be analyzed . [0078]

(Another embodiment of a method for analyzing inter-molecular interaction)

Next, another embodiment of the present method will be explained referring to FIG. 7 and FIG. 8.

As another embodiment, the present method may include (1) acquisition of the practical strength range of the magnetic particle MP, (2) acquisition of the saturated strength of a case of using the bonding pair elements of the analysis object, (3) setting of timing for adding the third molecule 3, and so on.

Also, the fundamental method and mechanism for reading a magnetic signal from the magnetic sensor MS may be general ones written in already reported literatures as described above. However, in general, with respect to the specification and information of the reagent and consumables selected and used by a user, such a case is common that the user himself /herself grasps them beforehand. They may be considered in reading a magnetic signal from the magnetic sensor MS. It is a matter of course that these specification and information are not to be limited to it, may be information described in a product and kit, and may be a value referred to in a literature. A simple example will be shown on the detection lower limit value and the detection upper limit value of the magnetic sensor MS and the magnetic particle MP, and the specification of the bonding pair elements (the first molecule 1 and the second molecule 2 ) .

[ 0079 ]

( 1 ) Acquisition of the practical strength range of the magnetic particle MP

FIG . 7 is a graph showing an example of the practical strength range of the magnetic particle MP which can be used from a detection lower limit value and a detection upper limit value of the magnetic sensor MS and the magnetic particle MP .

Magnetic signal strength 41 shown in FIG . 7 is strength MRp _max of the time when the magnetic particle MP bonds to the magnetic sensor MS at a maximum . MRp _max can be obtained easily only by making the magnetic particle MP of an excessive amount come into contact with the magnetic sensor MS .

Magnetic signal strength 45 is strength MRp _min of the time when the magnetic particle MP is not attached . MRp _min is the magnetic signal strength of the time when the magnetic signal is obtained only by the magnetic sensor MS .

The present method can set the detection lower limit value of the practical strength range of the magnetic particle MP to the magnetic signal strength 45 (MRp _min ) , and can set the detection upper limit value to the magnetic signal strength 41 (MRp _max ) .

[ 0080 ]

( 2 ) Acquisition of the saturated strength of a case of using the bonding pair elements of the analysis obj ect

Magnetic signal strength 42 shown in FIG . 7 is strength MRa _max of the time when the second molecule 2 to which the magnetic particle MP is labeled is made to bond at a maximum to the magnetic sensor MS to which the first molecule 1 is fixed at a maximum . Inevitably, the magnetic signal strength 42 becomes smaller than the magnetic signal strength 41 .

Similarly, magnetic signal strength 44 is strength MRa _min at which a magnetic signal is taken out while the magnetic sensor MS to which the first molecule 1 is fixed remains unchanged . In general , the magnetic signal strength 44 is larger than the magnetic signal strength 45 . [ 0081 ]

Also , when the magnetic sensor MS is one causing a solid phase reaction such as a 96 well microtiter plate frequently used in the immunological assay, the standard curve of the bonding pair elements ( the first molecule 1 and the second molecule 2 ) often becomes a sigmoid curve . In such a case , the lower limit of the linear range may be made MRa^ ( the magnetic signal strength 44 ) , and the upper limit may be made MRa _max ( the magnetic signal strength 42 ) . It is a matter of course that , when the standard curve does not become the sigmoid curve , namely when the standard curve is a straight line , the lower limit may be made MRp _min ( the magnetic signal strength 45 ) , and the upper limit may be made MRp _max ( the magnetic signal strength 41 ) .

On the whole , it is preferable that the magnetic signal range usable in inter-molecular interaction (practical strength range ) is made MRa _min ( the magnetic signal strength

44 ) to MRa _max ( the magnetic signal strength 42 ) . [ 0082 ] ( 3 ) Setting of timing for adding the third molecule 3

The present method can be roughly separated into two steps of before and after adding the third molecule 3 .

As shown in FIG . 7 , it is preferable that the timing t ₃ for adding the third molecule 3 in the third step S3 is set to such a range that a magnetic signal obtained does not go of f the dynamic range (practical strength range ) . When the third molecule 3 is added after going of f the dynamic range , it may possibly become hard to determine whether the reaction rate of the first molecule 1 and the second molecule 2 dropped by the influence of the third molecule 3 or boding of the first molecule 1 and the second molecule 2 saturated and the reaction rate dropped . When the third molecule 3 is added before going of f the dynamic range , such possibility can be reduced . The timing t ₃ for adding the third molecule 3 is , for example , preferable to be the time when the magnetic signal reaches (MRa _max -MRa _min ) /2 , and is more preferable to be before reaching (MRa _max -MRa _min ) /2 . By doing so , possibility that the determination becomes harder can be reduced more positively . However, when the timing t ₃ for adding the third molecule 3 is too early, it possibly becomes hard to correctly obtain the reaction rate parameter of the reaction of the first molecule 1 and the second molecule 2 . Therefore , usually ( in the standard mode ) , it is more preferable to set the timing t ₃ for adding the third molecule 3 to the time immediately before the magnetic signal reaches (MRa _max -MRa _min ) /2 .

[ 0083 ]

On the other hand, when a user wants to shorten the measuring time as much as possible ( shortest mode ) , the third molecule 3 can be added at freely-selected early timing t ₃ . In order to analyze inter-molecular interaction between the first molecule 1 and the second molecule 2 from temporal change of the first magnetic signal Sg1 , the timing t ₃ for adding the third molecule 3 may be set to the time of becoming the magnetic signal strength MRe _min that is minimally required for global fitting in Langmuir adsorption expression . This MRe _min can be investigated by a user beforehand, but may be set to the time of becoming approximately 10 times of the magnetic signal strength 45 (MRp _mim ) . [ 0084 ]

That is to say, when these five kinds of information (MRa _mim , MRp _mim , MRa _max , MRp _max , MRe _min ) on the consumables ( the magnetic sensor MS and the magnetic particle MP ) is grasped beforehand and is stored in a device that executes the present method and a desired condition is set , the third molecule 3 can be added automatically at the timing t ₃ when the real-time signal reaches the set time . Setting of them can be done as follows for example .

[ 0085 ] ■Input value: MRa _mim , MRp _mim , MRa _max , MRp _max , MRe _min ■Mode: standard/shortest ■In the standard mode: at the time when the real-time signal reaches (MRa _max -MRa _min ) / 2 (also, it is preferable to set to MRa _max <MRp _max and MRa _min >MRp _min ) ■In the shortest mode: at the time when the real-time signal reaches MRe _min (or 10 times of MRp _min ) [0086]

Also, the finishing timing for finishing to obtain temporal change of the second magnetic signal Sg2 after starting the third step S3 can be set freely. For example, the finishing timing can be set to the time of two times of the timing t ₃ . When the finishing time is set to this time, the present method and a device for executing the present method (a device for analyzing inter-molecular interaction 100 described below) can be finished automatically. The device for analyzing inter-molecular interaction 100 will be described below .

Thus, by the present method related to another embodiment, more complicated inter-molecular interaction related to biological matter can be analyzed easily.

[0087]

Example 2 related to another embodiment of the present method explained above will be shown below. Also, the contents of the present method are not limited by any means by this

Example 2. [0088]

(Example 2)

In Example 2, such a method was employed that the third molecule 3 was added after prescribed time elapsed after the reaction between the first molecule 1 and the second molecule 2 explained in [II] of Example 1 started.

In Example 2, first, the lower limit MRa _min and the upper limit MRa _max of the magnetic signal strength obtained by bonding the first molecule 1 and the second molecule 2 to each other were obtained.

By fixing pKA (the first molecule 1) used in the Example 1 to the surface of the GMR sensor by 1 pg, 10 pg, 100 pg, and 1 ng respectively, and the pKA-fixed magnetic sensor (the magnetic sensor device MSD) was manufactured. The pKA-fixed magnetic sensor was made to react within a well containing 400 μL of 1 nM magnetic nanoparticle-labeled quercetin solution (solution containing quercetin that was the second molecule 2) , and the magnetic signal after 20 minutes was obtained. Each of the fixing amount, the magnetic signal strength, and the result of the bonding rate constant obtained from the temporal change curve of them are shown in Table 3. Also, in Table 3, expresses that the bonding rate constant k _on could not be calculated or was not calculated. Also, " (-)MP" expresses that the magnetic particle MP is not used in the experiment system (namely exerts a role of the blank) . [0089] [Table 3]

[0090]

As shown in Table 3, when the pKA fixing amount is 0 pg, the magnetic signal strength was 11 ppm (MRa _min ) . When the pKA fixing amount is 1 ng, since the magnetic signal is deemed to have saturated, approximately 2,850 ppm of the magnetic signal strength at the 1 ng can be said to be MRp _max . However, with respect to MRa _min and MRa _max , since the magnetic signal strength is only 12 ppm even when the pKA fixing amount is 1 pg and the magnetic signal strength when the pKA fixing amount is 1 ng is not proportional to the fixing amount with respect to 100 pg, it cannot be said to be within the linear range. Therefore, this time, as the linear range, 118 ppm≈100 ppm was allocated for MRa _min and 1,250 ppm«1,200 ppm was allocated for MRa _max respectively .

Also, even when the bonding rate constant k _on was calculated as a reference value, a similar value was shown only in the linear range described above. [0091] Therefore, in the case of the standard mode, the timing t ₃ for adding the third molecule 3 becomes t ₃ = ( 1 , 200-100 ) /2=566 ppm. Actually, this calculation is executed within an electronic computer such as a personal computer (equivalent to a control unit 102 (FIG. 9, FIG. 10) described below) .

[0092]

There is shown in FIG. 8 an example of executing analysis of inter-molecular interaction of pKA (the first molecule 1) , quercetin (the second molecule 2) , and HSA (the third molecule 3) after inputting these values to the control unit 102. Also, FIG. 8 is a drawing showing an analysis result of inter- molecular interaction of the first molecule 1, the second molecule 2, and the third molecule 3 related to Example 2, and is a graph showing an example of temporal change of the magnetic signal strength.

[0093]

A solid line 51 shown in FIG. 8 shows an example of the temporal change curve obtained based on temporal change of the magnetic signal of a case the pKA-fixed magnetic sensor with 100 pg of the pKA fixing amount is made to react within a well containing 400 μL of 1 nM magnetic nanoparticle-labeled quercetin solution, and HSA (the third molecule 3) is added thereto so that the final density becomes 0.02%. Also, a broken line 52 shows an example of the temporal change curve obtained based on temporal change of the magnetic signal of a case of not adding the third molecule 3. [ 0094 ]

In FIG . 8 , the magnetic signal strength of the point where the solid line 51 and the broken line 52 branched was 571 ppm, and it was confirmed that HSA ( the third molecule 3 ) was added immediately after passing over 566 ppm having been set as described above . Also , in the case of the broken line 52 of not adding HSA, the calculation result of k _on was 1 . 9E+ 05 . Since this was of the same degree as that of the time of analysis by inter-molecular interaction based on the conventional di f ferent analysis method, it was confirmed that reliability was also secured . [ 0095 ]

Further, in the above , since the timing t ₃ for adding the third molecule 3 could be analyzed by the standard mode , it is considered that the analysis can be executed also by the shortest mode described above by changing setting . In the case of the shortest mode , it is considered that the analysis can be executed when the timing t ₃ is set either by MRe _min ( the magnetic signal strength minimally required for global fitting in Langmuir adsorption expression or by the magnetic signal strength) of 10 times of MRp _min namely 20 ppm from Table 3 . [ 0096 ]

From the above , it was confirmed that the timing t ₃ for adding the third molecule 3 could be set automatically by inputting information (MRa _mim , MRp _mim , MRa _max , MRp _max , MRe _min ) of consumables which could be grasped beforehand in general . Also , it was confirmed that , by doing so , a simple and highly precise device for analyzing inter-molecular interaction for biological matter could be materiali zed .

[ 0097 ]

[An embodiment of a device for analyzing inter-molecular interaction]

FIG . 9 is a schematic drawing showing an embodiment of a device for analyzing inter-molecular interaction 100 (will be hereinafter referred to as the present device 100 ) . In explanation below, a constituent element explained already is marked with a same reference sign, and detailed explanation thereof may be omitted . [ 0098 ]

As shown in FIG . 9 , the present device 100 includes a magnetism measuring unit 101 that allows the first molecule 1 , the second molecule 2 , and the third molecule 3 to react with each other and obtains a magnetic signal . Also , the present device 100 includes the control unit 102 that acquires the obtained magnetic signal with time to acquire temporal change thereof , executes various calculations , and executes various controls . [ 0099 ]

The magnetism measuring unit 101 uses the magnetic sensor MS with the first molecule 1 being fixed to the surface thereof , the second molecule 2 labeled by the magnetic particle MP, and the third molecule 3 , and obtains the first magnetic signal Sg1 and the second magnetic signal Sg2 in a manner described above . Also , the magnetism measuring unit 101 transmits the first magnetic signal Sg1 and the second magnetic signal Sg2 to the control unit 102 .

[ 0100 ]

The control unit 102 receives ( acquires ) the first magnetic signal Sg1 transmitted from the magnetism measuring unit 101 , and obtains the tendency T1 of temporal change of the first magnetic signal Sg1 . Also , the control unit 102 receives ( acquires ) the second magnetic signal Sg2 transmitted from the magnetism measuring unit 101 , and obtains the tendency T2 of temporal change of the second magnetic signal Sg2 . [ 0101 ]

Further, the control unit 102 compares the tendency T1 of temporal change of the first magnetic signal Sg1 and the tendency T2 of temporal change of the second magnetic signal Sg2 to each other, and, when these tendencies T1 , T2 of temporal change are di f ferent from each other ("NO" in FIG . 9 ) , analyzes that the third molecule 3 af fects inter-molecular interaction between the first molecule 1 and the second molecule 2 .

Also , when these tendencies T1 , T2 of temporal change are the same with each other ("YES" in FIG . 9 ) , the control unit 102 analyzes that the third molecule 3 does not af fect inter- molecular interaction between the first molecule 1 and the second molecule 2 .

[ 0102 ]

[Another embodiment of a device for analyzing inter-molecular interaction]

FIG . 10 is a schematic drawing explaining a block configuration and a flow of the present device 100 related to another embodiment . The present device 100 can have a configuration of being capable of automatically adding the third molecule 3 in addition to the configuration described above . As the configuration of being capable of automatically adding the third molecule 3 , an example of setting the timing ts described above can be employed .

The present device 100 employing a configuration of being capable of automatically adding the third molecule 3 will be hereinafter explained referring to FIG . 10 and in line with the contents of the processes . Also , explanation below is an example , and the present invention is not limited to it . [ 0103 ]

As shown in FIG . 10 , the present device 100 related to another embodiment includes the magnetism measuring unit 101 and the control unit 102 described above . First , the user fixes the first molecule 1 to the surface of the magnetic sensor MS in a manner described above , and manufactures the magnetic sensor device MSD ( S 101 ) . Also , the user labels the second molecule 2 by the magnetic particle MP ( S 102 ) . Further, the user arranges the magnetic sensor device MSD at a part of a prescribed component included in the magnetism measuring unit 101 namely the bottom part or the wall part of the well of the 96 well plate and the wall part of the flow cell for example .

[ 0104 ]

Further, the user inputs information of the consumables to the control unit 102 , and selects either of the standard mode or the shortest mode ( S 103 ) . With respect to the information of the consumables , MRa _mim , MRp _mim , MRa _max , MRp _max , MRe _min can be cited for example . When the information of the consumables and the mode are inputted, the control unit 102 stores them in a memory 104 .

Thereafter, the user operates a start button of the control unit 102 ( S 104 ) . When the start button is operated by the user, the control unit 102 transmits two start signals . [ 0105 ]

One of the start signals is transmitted toward the magnetism measuring unit 101 , and magnetism measurement is started . When magnetism measurement is started, the user adds the second molecule 2 to a prescribed portion where the magnetic sensor device MSD is arranged, the second molecule 2 being labeled by the magnetic particle MP ( S 105 ) . Thus , the first molecule 1 and the second molecule 2 bond to each other, and the inter-molecular interaction 12 occurs between them . When the first molecule 1 and the second molecule 2 come into contact with each other and a complex is formed, the first magnetic signal Sg1 detected on the magnetic sensor MS increases according to the amount of the complex . The magnetism measuring unit 101 measures the first magnetic signal Sg1 , and obtains temporal change thereof ( S 106 ) . The control unit 102 obtains this first magnetic signal Sg1 as a real-time signal MR1 , and obtains the tendency T1 of temporal change thereof . It is preferable that the real-time signal MR1 is transmitted to the control unit 102 on real-time . The control unit 102 stores temporal change of the first magnetic signal Sg1 ( the real-time signal MR1 ) in the memory 104 . [ 0106 ]

The other of the start signals allows determination of the timing t ₃ for adding the third molecule 3 to be executed within the control unit 102 ( S 107 ) . Determination of the timing t ₃ is executed according to either the standard mode or the shortest mode having been selected previously .

In the case of the standard mode , the control unit 102 acquires temporal change of the first magnetic signal Sg1 ( the real-time signal MR1 ) from the magnetism measuring unit 101 , refers to the information of the consumables stored in the memory 104 , and can set the timing t ₃ to the time when the real-time signal MR1 reaches (MRa _max -MRa _min ) /2 . Also , it is preferable to set to MRa _max <MRp _max and MRa _min RMRp _min .

In the case of the shortest mode , the control unit 102 refers to the information of the consumables stored in the memory 104 , and can set the timing t ₃ to the time when the real-time signal MR1 reaches MRe _min (or 10 times of MRp _min ) .

[0107]

When setting of the timing t ₃ finishes, the control unit 102 transmits an automatic injection trigger to a third molecule 3 adding tool 103 provided in the magnetism measuring unit 101. When the automatic injection trigger is received, the third molecule 3 adding tool 103 automatically adds the third molecule 3 at the timing t ₃ having been set to a solution where the first molecule 1 and the second molecule 2 bond to each other and the first magnetic signal Sg1 is generated (S108) . As the third molecule 3 adding tool 103, a conventional chemical feeder for example can be used. [0108]

Also, when setting of the timing t ₃ finishes, the control unit 102 allows the timing t ₃ to be stored in the memory 104. Next, the control unit 102 reads the timing t ₃ having been stored in the memory 104, and sets finishing time of magnetism measurement (S109) . The finishing time of magnetism measurement can be made two times of the timing t ₃ for example. [0109]

The magnetism measuring unit 101 executes magnetism measurement continuously even after adding the third molecule 3 (S110) , and the control unit 102 obtains temporal change of the second magnetic signal Sg2 as a real-time signal MR2. Similarly to the real-time signal MR1, it is preferable that also the real-time signal MR2 is transmitted to the control unit 102 on real-time . The control unit 102 allows temporal change of the second magnetic signal Sg2 ( the real-time signal MR2 ) to be stored in the memory 104 . As described above , for example , when the third molecule 3 reacts with and bonds to the second molecule 2 and so on and the inter-molecular interaction 13 occurs between the third molecule 3 and the second molecule 2 , the inter-molecular interaction 12 changes to inter-molecular interaction 12a by the influence of it . When the third molecule 3 exhibits a competitive nature , the tendency T2 of temporal change of the second magnetic signal Sg2 of the inter-molecular interaction 12a becomes smaller than the tendency T1 of temporal change of the first magnetic signal Sg1 of the inter-molecular interaction 12 . On the other hand, when the third molecule 3 exhibits a promotional nature , the tendency T2 of temporal change of the second magnetic signal Sg2 of the inter-molecular interaction 12a becomes larger than the tendency T1 of temporal change of the first magnetic signal Sg1 of the inter-molecular interaction 12 .

[ 0110 ]

When it becomes the finishing time of magnetism measurement , the control unit 102 transmits a signal of magnetism measurement finishing to the magnetism measuring unit 101 ( S 111) . When the signal of magnetism measurement finishing is received, the magnetism measuring unit 101 finishes magnetism measurement .

[ 0111 ] Also , when magnetism measurement finishes , the control unit 102 reads the tendency T1 of temporal change of the first magnetic signal Sg1 and the tendency T2 of temporal change of the second magnetic signal Sg2 from the memory 104 . Further, the control unit 102 compares the tendency T1 and the tendency T2 to each other ( S 112 ) . Also , when these tendencies T1 , T2 of temporal change are di f ferent from each other, the control unit 102 analyzes that the third molecule 3 af fects inter- molecular interaction between the first molecule 1 and the second molecule 2 .

On the other hand, when these tendencies T1 , T2 of temporal change are the same with each other, the control unit 102 analyzes that the third molecule 3 does not af fect inter- molecular interaction between the first molecule 1 and the second molecule 2 . [ 0112 ]

As described above , the present device 100 related to another embodiment can analyze not only inter-molecular interaction between the first molecule 1 and the second molecule 2 which is general as inter-molecular interaction but also influence of the third molecule 3 on inter-molecular interaction between these two molecules . In addition, the present device 100 can analyze more complicated inter- molecular interaction related to biological matter automatically and simply .

[ 0113 ] [ Program for analyzing inter-molecular interaction]

FIG . 11 is a flowchart showing a process flow of a program for analyzing inter-molecular interaction P related to the present embodiment (will be hereinafter referred to as "the present program P" ) along with a schematic configuration of the present device 100 .

The present program P allows a computer to execute a first procedure S i l and a second procedure S 12 described below in a method for analyzing inter-molecular interaction including the first step S1, the second step S2 , the third step S3 , and the fourth step S4 described above . The fourth step S4 is for comparing the tendency T1 of temporal change of the first magnetic signal Sg1 and the tendency T2 of temporal change of the second magnetic signal Sg2 to each other . As an example of a device using the present program P, explanation will be given citing the present device 100 shown in FIG . 11 . As shown in FIG . 11 , the present device 100 includes the magnetism measuring unit 101 and the control unit 102 ( computer ) .

[ 0114 ]

Also , for example , as shown in FIG . 11 , although the present program P can be configured to be installed in the memory 104 provided in the control unit 102 and to be cable of being started freely, the present invention is not limited to it . For example , it is possible that the present program P is installed on a freely-selected server computer on a net connected to be capable of two-way communication through an I/O port (not illustrated) provided in the control unit 102, and is started by the server computer. [0115] (First procedure Sil)

After the present program P is started, in the first procedure Sil, the control unit 102 is allowed to acquire the tendency T1 of temporal change of the first magnetic signal Sg1 obtained by bonding the first molecule 1 and the second molecule 2 to each other and the tendency T2 of temporal change of the second magnetic signal Sg2 obtained by adding the third molecule 3. Temporal change of the first magnetic signal Sg1 and temporal change of the second magnetic signal Sg2 can be stored once in the memory 104, and the control unit 102 can read them from the memory 104 from time to time. [0116] (Second procedure S12)

When the control unit 102 acquires the tendencies T1, T2, in the second procedure S12, the control unit 102 is allowed to compare these tendencies T1, T2 of temporal change to each other (T1=T2?) . Also, when these tendencies of temporal change are different from each other ("NO" in S12) , in the second procedure S12, the control unit 102 is allowed to analyze that the third molecule 3 affects inter-molecular interaction between the first molecule 1 and the second molecule 2. As described above, for example, when the third molecule 3 reacts with and bonds to the second molecule 2 and so on and the inter-molecular interaction 13 occurs between the third molecule 3 and the second molecule 2 , the inter-molecular interaction 12 changes to the inter-molecular interaction 12a by the influence of it . When the third molecule 3 exhibits a competitive nature , the tendency T2 of temporal change of the second magnetic signal Sg2 of the inter-molecular interaction 12a becomes smaller than the tendency T1 of temporal change of the first magnetic signal Sg1 of the inter-molecular interaction 12 . On the other hand, when the third molecule 3 exhibits a promotional nature , the tendency T2 of temporal change of the second magnetic signal Sg2 of the inter- molecular interaction 12a becomes larger than the tendency T1 of temporal change of the first magnetic signal Sg1 of the inter-molecular interaction 12 .

[ 0117 ]

On the other hand, in the second procedure S 12 , the control unit 102 is allowed to compare these tendencies T1 , T2 of temporal change to each other ( T1=T2 ? ) , and, when these tendencies of temporal change are the same with each other ("YES" in S 12 ) , is allowed to analyze that the third molecule 3 does not af fect the inter-molecular interaction described above . [ 0118 ] ( Summary)

As described above , the method, device , and program for analyzing inter-molecular interaction related to an embodiment of the present invention compare the tendency T1 of temporal change of the first magnetic signal Sg1 and the tendency T2 of temporal change of the second magnetic signal Sg2 to each other . Also , when these tendencies T1 , T2 of temporal change are di f ferent from each other, the method, device , and program for analyzing inter-molecular interaction related to an embodiment of the present invention analyze that the third molecule 3 af fects inter-molecular interaction between the first molecule 1 and the second molecule 2 . Further, when these tendencies T1 , T2 of temporal change are the same as each other, the method, device , and program for analyzing inter-molecular interaction related to an embodiment of the present invention analyze that the third molecule 3 does not af fect inter-molecular interaction between the first molecule 1 and the second molecule 2 . Thus , the method, device , and program for analyzing inter-molecular interaction related to an embodiment of the present invention can analyze not only inter-molecular interaction between the first molecule 1 and the second molecule 2 which is general as inter-molecular interaction but also influence of the third molecule 3 on inter-molecular interaction between these two molecules .

[ 0119 ]

Although the method, device , and program for analyzing inter-molecular interaction related to an embodiment of the present invention are explained in detail as described above , the gist of the present invention is not limited to it , and various modi fications are included .

Also , the embodiments described above are explained in detail to allow easy understanding of the present invention, and the present invention is not to be necessarily limited to one including all configurations explained . Further, a part of a configuration of an embodiment can be replaced by a configuration of other embodiments , and a configuration of an embodiment can be added with a configuration of other embodiments . Also , with respect to a part of a configuration of each embodiment , it is possible to ef fect addition, deletion, and replacement of other configurations .

Further, a part or all of each configuration, function, processing unit , processing means , controlling means , and the like described above may be achieved by hardware such as being designed by an integrated circuit for example . Also , each configuration, function, and the like described above may be achieved by software by that a processor interprets and executes programs achieving each function . Information of a program, a table , a file , and the like achieving each function can be placed in a storage device such as a memory, a hard disk, and a SSD ( Solid State Drive ) or in a recording medium such as an IC card, a SD card, and a DVD .

Also , a control line and information line are shown for those considered to be necessary for explanation, and are not necessarily limited to show all control lines and information lines necessary for a product . In fact , it can be considered that almost all configurations are connected to each other.

[List of Reference Signs]

[0120]

51. . . First step

S2. . . Second step

53. . . Third step

54... Fourth step

1. . . First molecule

2. . . Second molecule

3... Third molecule

MS... Magnetic sensor

MP... Magnetic particle

MSD... Magnetic sensor device Sg1. . . First magnetic signal

Sg2. . . Second magnetic signal

Ti, T2. . . Tendency

100... Device for analyzing inter-molecular interaction (present device)

101... Magnetism measuring unit

102... Control unit

P... Program for analyzing inter-molecular interaction (present program)

511... First procedure

S12... Second procedure