Anti-peg antibody–binding material and detection method for Anti-peg antibody

Abstract

Provided is an anti-PEG antibody–binding material that makes it possible to detect an anti-PEG antibody. According to the embodiments, an anti-PEG antibody–binding material includes a cellulose oligomer represented by general formula (1). In formula (1), n represents a number that is at least 6 but no greater than 16, m represents a number that is at least 3, and A represents a C1–24 divalent organic group that links the oxygen atom bonded to the anomeric carbon at the reducing end of the cellulose oligomer and the polyethylene glycol chain and includes at least one type of heteroatom selected from the group that consists of nitrogen atoms, oxygen atoms, and sulfur atoms.

Description

Anti-PEG antibody binding material and method for detecting anti-PEG antibodies

[0001] The present invention relates to an anti-PEG antibody binding material and a method for detecting an anti-PEG antibody using the same.

[0002] Modification of the surface of proteins, liposomes, micelles, etc. with PEG (polyethylene glycol) improves the blood circulation of the modified substance and reduces immunogenicity. For this reason, many PEG-modified pharmaceuticals are currently in clinical use.

[0003] However, it has been revealed that PEG modification induces the secretion of antibodies against PEG (anti-PEG antibodies). If a PEG-modified drug is administered while anti-PEG antibodies are present in the blood, the PEG-modified drug may be removed from the blood (ABC phenomenon), resulting in reduced blood retention and side effects. Therefore, there is a need for highly sensitive detection of anti-PEG antibodies present in the blood in order to evaluate the induction of anti-PEG antibodies and their effect on the therapeutic efficacy of PEG-modified drugs.

[0004] Anti-PEG antibodies include backbone-selective anti-PEG antibodies and methoxy-terminated PEG-selective anti-PEG antibodies. Backbone-selective anti-PEG antibodies recognize the repeating oxyethylene structure that forms the backbone of the PEG structure and specifically bind to the PEG chain. Methoxy-terminated PEG-selective anti-PEG antibodies are antibodies that have high selectivity for methoxy-terminated PEG, i.e., they specifically bind to PEG chains having a methoxy group at the end.

[0005] As a technology relating to biosensors containing PEG chains, for example, Patent Document 1 discloses a biosensor containing PEG-modified nanoparticles in which functional groups or functional moieties are attached to metal (oxide) microparticles or semiconductor microparticles via two or more PEG chains.

[0006] Furthermore, Patent Documents 2 and 3 disclose biosensors containing hydrophilic particles, each of which includes water-insoluble particles, a primer layer containing polysiloxane disposed on the surface of the particles, and a hydrophilic polymer layer disposed on the primer layer, the hydrophilic polymer layer being composed of PEG chains to which antibodies are bound and PEG chains to which inactive groups are bound.

[0007] JP 2005-180921 A JP 2020-143964 A JP 2021-012089 A

[0008] In the course of research aimed at detecting anti-PEG antibodies present in the blood with high sensitivity, the inventors discovered that certain cellulose oligomers bearing PEG chains specifically bind to anti-PEG antibodies.

[0009] The embodiments of the present invention have been made in view of the above points, and have an object to provide an anti-PEG antibody-binding material capable of binding to an anti-PEG antibody, and a method for detecting an anti-PEG antibody using the same.

[0010] The present invention includes the following embodiments: [1] An anti-PEG antibody binding material comprising a cellulose oligomer represented by general formula (1). (In formula (1), n ​​represents the average degree of polymerization and is a number from 6 to 16; m represents the average degree of polymerization and is a number not less than 3; and A represents a divalent organic group connecting the oxygen atom bonded to the anomeric carbon at the reducing end of the cellulose oligomer and the polyethylene glycol chain, the divalent organic group having 1 to 24 carbon atoms and containing at least one heteroatom selected from the group consisting of a nitrogen atom, an oxygen atom, and a sulfur atom.) [2] The anti-PEG antibody binding material according to [1], wherein A in formula (1) represents a divalent organic group having 2 to 10 carbon atoms and containing at least one heteroatom selected from the group consisting of a triazole group, an amide group, and a thioether group. [3] The anti-PEG antibody binding material according to [1], wherein A in formula (1) represents where R １ represents a hydrocarbon group having 1 to 10 carbon atoms, and R ２[4] The anti-PEG antibody binding material according to any one of [1] to [3], wherein m in formula (1) represents a number of 10 or more.

[0011] [5] A biosensor comprising the anti-PEG antibody-binding material according to any one of [1] to [4]. [6] An adsorbent comprising the anti-PEG antibody-binding material according to any one of [1] to [4].

[0012] [7] A method for detecting an anti-PEG antibody, comprising: incubating a cellulose oligomer represented by the above general formula (1) with a test specimen suspected of containing the anti-PEG antibody under conditions that allow binding of the cellulose oligomer with the anti-PEG antibody; incubating the cellulose oligomer after incubation with the test specimen with a sample containing the secondary antibody under conditions that allow binding of the secondary antibody to the anti-PEG antibody; and using the cellulose oligomer after incubation with the sample to detect the anti-PEG antibody contained in the test specimen with the secondary antibody as a probe.

[0013] According to an embodiment of the present invention, there is provided an anti-PEG antibody-binding material capable of binding to an anti-PEG antibody, which can be used, for example, in a biosensor for detecting an anti-PEG antibody or as an adsorbent for adsorbing an anti-PEG antibody.

[0014] 1 is a conceptual diagram of a cellulose nanosheet (cellulose oligomer assembly) made of cellulose oligomers constituting an anti-PEG antibody binding material according to one embodiment. 2 is a reaction scheme of a PEG-introduced cellulose oligomer according to one embodiment. 3 is an explanatory diagram showing a method for detecting an anti-PEG antibody according to one embodiment. 4 is a diagram showing mass spectra of the cellulose oligomer assemblies obtained in Synthesis Examples 1 to 4. 5 is a diagram showing ATR / FT-IR spectra of the cellulose oligomer assemblies obtained in Synthesis Examples 1 to 4. 6 is a graph showing the results (absorbance at 490 nm) of binding of an anti-PEG antibody (PEG-B-47) to each cellulose oligomer assembly in Test Example 1. 7 is a graph showing the results (absorbance at 490 nm) of binding of an anti-PEG antibody (15-2b) to each cellulose oligomer assembly in Test Example 2. 8 is a graph showing the results (absorbance at 490 nm) of binding of an anti-PEG antibody (6.3) to each cellulose oligomer assembly in Test Example 3.

[0015] The anti-PEG antibody binding material according to this embodiment contains a cellulose oligomer represented by general formula (1) (hereinafter referred to as a PEG-introduced cellulose oligomer). A PEG-introduced cellulose oligomer is a cellulose oligomer having a structure in which glucose units are linked by β-1,4-glucosidic bonds, with a substituent containing a polyethylene glycol chain (PEG chain) introduced to the anomeric carbon at its reducing end. More specifically, a PEG chain having a methoxy group at its end is bound to the oxygen atom bound to the anomeric carbon via linking group A. By incorporating linking group A in this manner, PEG chains of various chain lengths can be introduced into the cellulose oligomer.

[0016]

[0017] In formula (1), n ​​is a number from 6 to 16, m is a number of 3 or more, and A is a divalent organic group. The wavy line in formula (1) indicates that the anomeric configuration at the reducing end is an α-form, a β-form, or a mixture of α- and β-forms.

[0018] The above n represents the average degree of polymerization (DP) of the cellulose oligomer and is 6 or more and 16 or less. n may be 6 to 14, 6.5 to 13, or 7 to 12. The average degree of polymerization of the cellulose oligomer is a weighted average (number-average degree of polymerization) of the degrees of polymerization according to the molecular ratio of cellulose oligomers with different degrees of polymerization. The degree of polymerization of each PEG-introduced cellulose oligomer is not particularly limited, but may be, for example, 4 to 20, 5 to 16, or 6 to 13.

[0019] The above m represents the average degree of polymerization (DP) of the PEG chain and is 3 or more. m can be set depending on the selectivity of the anti-PEG antibody to be bound; for example, by setting m to 10 or more, backbone-selective anti-PEG antibodies can be bound more easily. The upper limit of m is not particularly limited, but is preferably 500 or less. In one embodiment, m may be 3 to 500, 10 to 250, 16 to 150, 18 to 100, or 20 to 50. The average degree of polymerization of the PEG chain is a weighted average of the degrees of polymerization according to the molecular ratio of PEG chains with different degrees of polymerization. Note that when there is no distribution in the molecular weight of the PEG chain (i.e., when the polydispersity is 1), m simply takes on an integer value as the degree of polymerization.

[0020] The above A is an oxygen atom bonded to the anomeric carbon at the reducing end of the cellulose oligomer and a PEG chain (i.e., (CH ２ CH ２ O) ｍ ) and represents a divalent organic group (linking group) that contains at least one heteroatom selected from the group consisting of a nitrogen atom, an oxygen atom, and a sulfur atom and has 1 to 24 carbon atoms.

[0021] The number of carbon atoms in the linking group A is preferably 1 to 20, more preferably 2 to 15, and even more preferably 2 to 10. Examples of groups containing a heteroatom contained in the linking group A include a triazole group, an amide group, a thioether group, a maleimide group, an ether group, and an ester group, and the cellulose oligomer and the PEG chain are linked via these heteroatom-containing groups. Among these, from the viewpoints of ease of synthesis, bond stability, and the like, it is preferable that the linking group A contains a triazole group, an amide group, or a thioether group.

[0022] In one embodiment, A in formula (1) preferably represents a divalent organic group having 2 to 10 carbon atoms and containing at least one selected from the group consisting of a triazole group, an amide group, and a thioether group.

[0023] More specifically, A in formula (1) is It is preferred that

[0024] Here, R １ represents a hydrocarbon group (divalent hydrocarbon group) having 1 to 10 carbon atoms, and is bonded to the oxygen atom bonded to the anomeric carbon at the reducing end. １ is preferably a linear or branched alkanediyl group, more preferably a linear alkanediyl group. １ The hydrocarbon group preferably has 1 to 6 carbon atoms, more preferably 1 to 3 carbon atoms.

[0025] R ２ represents a single bond or a hydrocarbon group having 1 to 10 carbon atoms, and is bonded to the PEG chain. ２ is preferably a single bond or a linear or branched alkanediyl group, and more preferably a single bond (i.e., a direct bond). ２ The hydrocarbon group preferably has 1 to 6 carbon atoms, more preferably 1 to 3 carbon atoms.

[0026] In one embodiment, the PEG-introduced cellulose oligomer is preferably represented by the following general formula (2): In this case, the anomeric position at the reducing end has a β-configuration. １ and R２ is as above.

[0027] The PEG-introduced cellulose oligomer may be contained in a cellulose oligomer assembly having a cellulose type II crystalline structure. The cellulose oligomer assembly may have a sheet-like structure (cellulose nanosheet). Here, the sheet-like structure is a concept that encompasses a ribbon-like structure (cellulose nanoribbon) as shown in Figure 1.

[0028] While naturally occurring cellulose chains have a parallel-aligned cellulose type I crystal structure, artificially synthesized cellulose oligomers generally form the thermodynamically stable cellulose type II crystal structure. In this case, the substituents at the ends of the cellulose chains do not affect the crystal shape, and cellulose oligomers with substituents align in the thickness direction of the cellulose nanosheet to form lamellar crystals, with the substituents exposed on the surface of the sheet.

[0029] In detail, as one embodiment, a structure in which a substituent containing a PEG chain has been introduced into some of the cellulose oligomers constituting a cellulose nanosheet is shown in Figure 1. In Figure 1, the PEG-introduced cellulose oligomer is shown with an arrow pointing from the reducing end to the non-reducing end, as noted below the above formula (1), and the substituent containing the PEG chain is shown with a wavy line. Furthermore, for a cellulose oligomer without a PEG chain introduced, the cellulose chain is shown with an arrow pointing from the reducing end to the non-reducing end. As shown in the cross-sectional structure in Figure 1, the cellulose oligomer containing the PEG-introduced cellulose oligomer is arranged in the film thickness direction so that the directions of the arrows alternate, and the substituent containing the PEG chain is exposed on the sheet surface.

[0030] The synthesis method for PEG-introduced cellulose oligomers is not particularly limited, but they may be synthesized as follows. That is, for example, a cellulose oligomer having a functional group introduced at the anomeric position of the reducing end is used to prepare a cellulose nanosheet from the functionalized cellulose oligomer. By reacting a compound having a PEG chain (PEG compound) with the functional group exposed on the surface of the cellulose nanosheet, the PEG chain can be introduced into the cellulose oligomer via the linking group A. In this way, by using a cellulose nanosheet having a functional group exposed on the sheet surface and introducing a PEG chain by reacting with the functional group, it is possible to introduce a PEG chain with a long chain length. Therefore, binding with an anti-PEG antibody that specifically binds to the long PEG chain becomes possible.

[0031] Examples of the reaction between the functional group and a compound having a PEG chain include a click reaction between an azide group and an alkynyl group, an amidation reaction between a carboxy group and an amino group, an esterification reaction between a carboxy group and a hydroxy group, a thiol-ene reaction between a thiol group and a carbon-carbon double bond, and a reaction between a thiol group and an organic halide.

[0032] FIG. 2 shows a reaction scheme for PEG-introduced cellulose oligomers according to one embodiment. Using a cellulose nanosheet with ethynyl groups exposed on the surface as functional groups, a click reaction is carried out with a PEG compound having an azide group using a metal catalyst such as a copper catalyst. This introduces a PEG chain into the cellulose oligomer via linking group A containing a triazole group. This is represented by the following chemical formula (3). In reaction formula (3), a click reaction is carried out between a cellulose oligomer (CEL-C≡CH) having a propargyl group and methoxypolyethylene glycol azide (mPEG azide).

[0033] In reaction formula (3), the functional group of the cellulose oligomer is an alkynyl group, and the functional group of the PEG compound is an azide group, but this may be reversed. That is, as shown in reaction formula (4) below, a click reaction may be carried out between a cellulose oligomer having an azide group as a functional group and a PEG compound having an ethynyl group.

[0034] In another embodiment, as shown in the following reaction formula (5), a cellulose oligomer having an amino group as a functional group and a PEG compound having a carboxy group may be subjected to an amidation reaction, thereby introducing a PEG chain into the cellulose oligomer via a linking group A containing an amide group.

[0035] In reaction formula (5), the functional group of the cellulose oligomer is an amino group, and the functional group of the PEG compound is a carboxy group, but this may be reversed. That is, as shown in the following reaction formula (6), a cellulose oligomer having a carboxy group as a functional group may be subjected to an amidation reaction with a PEG compound having an amino group.

[0036] In another embodiment, as shown in the following reaction formula (7), a cellulose oligomer having a thiol group as a functional group may be subjected to a thiol-ene reaction with a PEG compound having a maleimide group, thereby introducing a PEG chain into the cellulose oligomer via a linking group A containing a thioether group.

[0037] In reaction formula (7), the functional group of the cellulose oligomer is a thiol group, and the functional group of the PEG compound is a maleimide group, but this may be reversed. That is, as shown in reaction formula (8) below, a cellulose oligomer having a maleimide group as a functional group may be subjected to a thiol-ene reaction with a PEG compound having a thiol group.

[0038] In another embodiment, as shown in the following reaction formula (9), a cellulose oligomer having a thiol group as a functional group may be reacted with a PEG compound having a halogen atom, thereby introducing a PEG chain into the cellulose oligomer via a linking group A containing a thioether group.

[0039] The anti-PEG antibody-binding material according to this embodiment contains a PEG-introduced cellulose oligomer, preferably contained as a cellulose oligomer assembly such as a cellulose nanosheet. That is, in one embodiment, the anti-PEG antibody-binding material preferably contains a cellulose oligomer assembly containing a PEG-introduced cellulose oligomer. The cellulose oligomer assembly may consist solely of a PEG-introduced cellulose oligomer, or may contain a cellulose oligomer without a PEG chain introduced therein in addition to the PEG-introduced cellulose oligomer.

[0040] When a cellulose oligomer assembly comprising functionalized cellulose oligomers is used and reacted with a PEG compound as described above, the anti-PEG antibody binding material may comprise a cellulose oligomer assembly containing a PEG-introduced cellulose oligomer and a functionalized cellulose oligomer. In the cellulose oligomer assembly, the proportion of PEG-introduced cellulose oligomers to all cellulose oligomers constituting the assembly (PEG incorporation rate) is not particularly limited and may be, for example, 1 mol % or more, preferably 5 to 80 mol %, more preferably 7 to 60 mol %, and even more preferably 10 to 50 mol %.

[0041] The PEG-introduced cellulose oligomer binds to anti-PEG antibodies such as backbone-selective anti-PEG antibodies and methoxy-terminated PEG-selective anti-PEG antibodies, but does not bind to secondary antibodies used as probes or other proteins such as BSA (bovine serum albumin) and lysozyme, and can therefore be used as an anti-PEG antibody-binding material capable of detecting or adsorbing anti-PEG antibodies.

[0042] Examples of anti-PEG antibodies to which PEG-introduced cellulose oligomers can be bound include backbone-selective anti-PEG antibodies and / or methoxy-terminated PEG-selective anti-PEG antibodies. The class of anti-PEG antibodies is not particularly limited, and examples include IgG, IgG1, IgG2b, and IgM.

[0043] A backbone-selective anti-PEG antibody is a monoclonal antibody that recognizes the repeating oxyethylene unit that constitutes the backbone of the PEG structure and specifically binds to the PEG chain. The molecular weight of the PEG chain that can specifically bind varies depending on the type of anti-PEG antibody. Therefore, the average degree of polymerization m of the PEG chain in the above formula (1) can be set depending on the type of anti-PEG antibody to be detected or adsorbed.

[0044] For example, the anti-PEG antibody "6.3" specifically binds to the backbone of a PEG chain of 750 Da (DP=16) or more, and therefore, when attempting to detect or adsorb this, the average degree of polymerization m of the PEG chain in formula (1) can be set to 16 or more. For example, the anti-PEG antibody "AGP4" specifically binds to PEG chains of 2000 Da (DP=45) or more, and therefore, when attempting to detect or adsorb this, the average degree of polymerization m of the PEG chain in formula (1) can be set to 45 or more.

[0045] A methoxy-terminated PEG-selective anti-PEG antibody is a monoclonal antibody that recognizes and specifically binds to PEG chains having a methoxy group at their terminus. A methoxy-terminated PEG-selective anti-PEG antibody may also bind to PEG chains having a more hydrophobic ethoxy or butoxy group at their terminus. The molecular weight of the PEG chain that can specifically bind varies depending on the type of anti-PEG antibody. Therefore, the average degree of polymerization m of the PEG chains in the above formula (1) can be set depending on the type of anti-PEG antibody to be detected or adsorbed.

[0046] For example, the anti-PEG antibody "15-2b" specifically binds to PEG chains of 560 Da (DP=12) or more having a methoxy group at the end, and therefore, when attempting to detect or adsorb this, the average degree of polymerization m of the PEG chains in formula (1) can be set to 12 or more. For example, the anti-PEG antibody "PEG-B-47" specifically binds to PEG chains of 560 Da (DP=12) or more having a methoxy group at the end, and therefore, when attempting to detect or adsorb this, the average degree of polymerization m of the PEG chains in formula (1) can be set to 3 or more.

[0047] The anti-PEG antibody-binding material according to the embodiment contains a PEG-introduced cellulose oligomer and is therefore capable of recognizing and binding to an anti-PEG antibody, and can therefore be used, for example, as a biosensor for detecting an anti-PEG antibody or an adsorbent for adsorbing an anti-PEG antibody.

[0048] The anti-PEG antibody binding material may consist solely of a PEG-introduced cellulose oligomer or a cellulose oligomer aggregate containing the same, or may contain other components such as proteins, peptides, nucleic acids, lipids, sugars, cells, amino acids, metal ions, organic solvents, etc., within the scope that does not impair its effect.

[0049] The PEG-introduced cellulose oligomer or cellulose oligomer aggregate is a substance that does not dissolve in water, and when prepared by the above synthesis method, it is obtained as an aqueous dispersion of cellulose nanosheets containing the PEG-introduced cellulose oligomer. The anti-PEG antibody-binding material according to one embodiment may be a dry powder obtained by removing water from the aqueous dispersion. A powdered anti-PEG antibody-binding material may be prepared using the dry powder as is, or by powder-mixing with other components such as proteins. The powdered anti-PEG antibody-binding material can be easily dispersed in water by adding water when in use, thereby preparing an aqueous dispersion.

[0050] An anti-PEG antibody binding material according to one embodiment may be an aqueous dispersion in which a PEG-introduced cellulose oligomer or cellulose oligomer assembly is dispersed in water. In this case, a buffer solution prepared by adding a buffer to water may be used as the dispersion medium. In other words, the aqueous dispersion may be a dispersion of a PEG-introduced cellulose oligomer or cellulose oligomer assembly in a buffer solution. The content (concentration) of the PEG-introduced cellulose oligomer in the aqueous dispersion is not particularly limited and may be, for example, 0.01 to 5% (w / v) or 0.03 to 0.3% (w / v).

[0051] In this specification, "% (w / v)" means a mass volume percent concentration, which is the mass (g) of the target substance contained in a volume of 100 mL.

[0052] The buffer is not particularly limited, and examples thereof include buffers that constitute phosphate buffered saline (PBS), 2-morpholinoethanesulfonic acid (MES) buffer, trishydroxymethylaminomethane (Tris) buffer, 3-morpholinopropanesulfonic acid (MOPS) buffer, and 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) buffer.

[0053] An anti-PEG antibody-binding material according to one embodiment may be a PEG-introduced cellulose oligomer or cellulose oligomer assembly immobilized on the surface of a substrate such as plastic or glass.

[0054] In one embodiment, a biosensor comprising the anti-PEG antibody binding material is used to perform a bioassay to detect and quantify anti-PEG antibodies, and the term biosensor refers to each element or assembly or combination of elements used to perform such a bioassay.

[0055] Therefore, the biosensor may be composed solely of a powder of PEG-introduced cellulose oligomer or cellulose oligomer aggregate, or may be composed solely of an aqueous dispersion of PEG-introduced cellulose oligomer or cellulose oligomer aggregate. Alternatively, the biosensor may be a kit combining a powder or aqueous dispersion of the PEG-introduced cellulose oligomer or cellulose oligomer aggregate with a secondary antibody as a probe (i.e., an anti-PEG antibody detection kit). Furthermore, if the secondary antibody is an enzyme-labeled antibody, the biosensor may be an anti-PEG antibody detection kit combining a substrate corresponding to the enzyme.

[0056] The biosensor may contain albumin such as BSA, and albumin is preferably added when incubating the PEG-introduced cellulose oligomer or cellulose oligomer assembly with the test specimen. In this case, albumin may be included in the anti-PEG antibody binding material or may be included in the anti-PEG antibody detection kit separately from the anti-PEG antibody binding material. When albumin is included in the anti-PEG antibody binding material, the albumin may be in powder form mixed with the PEG-introduced cellulose oligomer or cellulose oligomer assembly, or the albumin may be dissolved in an aqueous dispersion. The ratio of PEG-introduced cellulose oligomer to albumin is not particularly limited, and may be, for example, a PEG-introduced cellulose oligomer / albumin mass ratio of 0.002 / 1 to 0.3 / 1 or 0.01 / 1 to 0.1 / 1. Note that when the test specimen contains albumin, such as blood, additional albumin may or may not be added.

[0057] A method for detecting an anti-PEG antibody according to one embodiment comprises the following steps: (1) incubating a PEG-introduced cellulose oligomer with a test specimen under conditions that allow binding of the anti-PEG antibody, (2) incubating the PEG-introduced cellulose oligomer after incubation with the test specimen with a sample containing a secondary antibody under conditions that allow binding of the secondary antibody, and (3) using the PEG-introduced cellulose oligomer after incubation with the sample as a probe to detect the anti-PEG antibody contained in the test specimen with the secondary antibody.

[0058] In step (1), a test sample is added to a container together with an aqueous dispersion of PEG-introduced cellulose oligomer as the anti-PEG antibody-binding material, mixed and dispersed, and then incubated. The aqueous dispersion is preferably an aqueous dispersion of a cellulose oligomer aggregate, which is an aggregate containing PEG-introduced cellulose oligomer.

[0059] The analyte is a substance to be tested that is suspected of containing anti-PEG antibodies. For example, when detecting anti-PEG antibodies present in blood to evaluate the influence of the anti-PEG antibody induction or the therapeutic effect of a PEG-modified pharmaceutical, the analyte may be blood or a diluted version of the blood.

[0060] In step (1), as described above, albumin such as BSA may be added to improve the detection sensitivity of the anti-PEG antibody. In this case, the concentration of albumin in the system (the albumin concentration excluding albumin originally contained in the test sample) is not particularly limited, but is preferably, for example, 0.05 to 10% by mass, and more preferably 0.5 to 5% by mass.

[0061] The incubation conditions in step (1) are not particularly limited as long as they allow binding of the PEG-introduced cellulose oligomer to the anti-PEG antibody, and may be, for example, 4 to 60°C and 1 minute to 2 hours. The concentration of the PEG-introduced cellulose oligomer in the system is not particularly limited, and may be, for example, 0.001 to 1% (w / v) or 0.01 to 0.1% (w / v).

[0062] In step (1), the anti-PEG antibody to be detected is bound to the PEG-introduced cellulose oligomer (see Figure 3).The PEG-introduced cellulose oligomer to which the anti-PEG antibody is bound is then separated from the specimen by centrifugation and washing.

[0063] In step (2), a sample containing a secondary antibody is added to a PEG-introduced cellulose oligomer (preferably a cellulose oligomer assembly containing the same) to which an anti-PEG antibody has been bound, and the mixture is mixed and dispersed, followed by incubation.

[0064] The sample containing the secondary antibody may be an aqueous solution of the secondary antibody. The secondary antibody is not particularly limited as long as it can be used as a probe by specifically binding to the anti-PEG antibody, and examples thereof include antibodies labeled with an enzyme, a radioisotope, a fluorescent dye, or the like.

[0065] The incubation conditions in step (2) are not particularly limited as long as they allow binding between the anti-PEG antibody and the secondary antibody, and may be, for example, 4 to 60°C and 1 minute to 2 hours. The concentration of the secondary antibody in the system is not particularly limited, and may be, for example, 100 to 1000 ng / mL.

[0066] In step (2), a secondary antibody is bound as a probe to the anti-PEG antibody bound to the PEG-introduced cellulose oligomer (see Figure 3).The PEG-introduced cellulose oligomer to which the secondary antibody is bound (preferably a cellulose oligomer assembly containing the PEG-introduced cellulose oligomer) is then separated from the sample containing the secondary antibody by centrifugation and washing.

[0067] In step (3), a PEG-introduced cellulose oligomer to which a secondary antibody is bound is used to detect and quantify the anti-PEG antibody contained in the sample using the secondary antibody as a probe.

[0068] For example, when an enzyme-labeled antibody is used as the secondary antibody, an enzyme reaction is carried out by adding a substrate corresponding to the enzyme to the cellulose oligomer assembly to which the secondary antibody is bound, as shown in Figure 3. The resulting product pigment is detected and quantified by absorbance (color development), fluorescence, chemiluminescence, or the like.

[0069] In another embodiment, an adsorbent comprising the anti-PEG antibody-binding material can be used to remove or purify the anti-PEG antibody by adsorbing the anti-PEG antibody.

[0070] The adsorbent may be composed of a powder or aqueous dispersion of a PEG-introduced cellulose oligomer or a cellulose oligomer aggregate containing the same, and these may contain other optional components.

[0071] When an adsorbent is used to remove anti-PEG antibodies, for example, the adsorbent is added to a liquid containing the anti-PEG antibodies, and the anti-PEG antibodies are brought into contact with the PEG-introduced cellulose oligomer to bind to each other. The anti-PEG antibodies can then be removed from the liquid by centrifuging or other methods to remove the adsorbent with the adsorbed anti-PEG antibodies.

[0072] When an adsorbent is used to purify an anti-PEG antibody, for example, the adsorbent is added to a liquid containing the anti-PEG antibody and impurities, and the anti-PEG antibody is brought into contact with a PEG-introduced cellulose oligomer, thereby binding the two. The adsorbent with the adsorbed anti-PEG antibody is then separated and recovered from the liquid by centrifugation or other methods. The recovered adsorbent can be treated with an acid, for example, to release the bond between the anti-PEG antibody and the PEG-introduced cellulose oligomer, allowing the anti-PEG antibody to be separated and recovered from the adsorbent, resulting in a highly pure anti-PEG antibody.

[0073] The present invention will be further explained below with reference to examples, but is not limited to these.

[0074] [Enzymatic synthesis of cellulose oligomers bearing propargyl groups] A 1M aqueous solution of α-glucose-1-phosphate (αG1P), a 1M 2-[4-(2-hydroxyethyl)-1-piperazinyl]ethanesulfonic acid (HEPES) buffer (pH 7.5), a 100 μM EDTA solution, and a 1M aqueous solution of 2-propynyl-β-D-glucoside (Glc-C≡CH) were mixed in predetermined amounts, and ultrapure water was added to prepare a solution containing 200 mM αG1P, 500 mM HEPES, 50 μM EDTA, and 50 mM Glc-C≡CH. Here, "M" stands for "mol / L." Cellodextrin phosphorylase (CDP) was added to the resulting solution at 1 U / mL, followed by incubation at 60°C for 3 days. The liquid containing the precipitated product was centrifuged (20,400 × g, 5 minutes, 4°C), and the supernatant was removed. Ultrapure water was then added to redisperse the product, and the centrifugal separation (under the same conditions) was repeated five times to obtain a cellulose oligomer having propargyl groups (CEL-C≡CH).

[0075] The above 2-propynyl-β-D-glucoside was synthesized as follows: Tetra-O-acetyl-2-propynyl-β-D-glucoside was synthesized according to a previous report (H.B. Mereyala et al., Carbohydr. Polym. 1998, 307, 351-354). Next, 0.97 g of tetra-O-acetyl-propargyl-β-D-glucoside was reacted with 0.034 g of sodium methoxide in the presence of 25 mL of methanol and 25 mL of tetrahydrofuran at 25°C for at least 12 hours, and the reaction mixture was then purified by silica gel column chromatography to obtain 2-propynyl-β-D-glucoside.

[0076] The fact that the product obtained by enzymatic synthesis was a cellulose oligomer having a propargyl group at one end was confirmed by using a proton nuclear magnetic resonance (NMR) spectrometer (AVANCE III HD500 (Bruker Biospin, magnetic field strength: 500 MHz, number of accumulations: 16)) using a sample prepared by dissolving the product in a 4% deuterium oxide heavy water solution at a concentration of 2% by mass or more.

[0077] The mass spectrum of the product was obtained by matrix-assisted laser desorption / ionization time-of-flight mass spectrometry (MALDI-TOF-MS), which revealed that the average degree of polymerization of the cellulose oligomer was 7.5 and the polydispersity index (PDI) was 1.02. The measurement conditions for MALDI-TOF-MS and the calculation methods for the average degree of polymerization and polydispersity index (PDI) are as follows:

[0078] Measurement was performed using a MALDI-TOF MS device (AXIMA-performance (Shimadzu Corporation)) under the following conditions: acceleration voltage 20 kV, linear (positive) ion mode, mass range 1-4000, max laser rap rate 10, power 100, profiles 100, shots 2, ion gate blank 500 Da, pulsed extraction optimized at 1500 Da. The average degree of polymerization and polydispersity index (PDI) were calculated based on the integrated value of the peak of a sodium or potassium adduct of cellulose oligomer (CEL-C≡CH) having a degree of polymerization of 6 to 11 and a propargyl group at one end.

[0079] [Introduction of PEG chains into cellulose oligomers] A click reaction was carried out between a cellulose oligomer having a propargyl group (CEL-C≡CH) and methoxypolyethylene glycol azide (mPEG azide) according to the reaction scheme shown in Figure 2 and the reaction shown in Reaction Formula (3) above. Four types of PEG-introduced cellulose oligomers with different PEG incorporation rates were prepared according to Synthesis Examples 1 to 4 below.

[0080] Synthesis Example 1 A CEL-C≡CH aqueous dispersion, 40 mM methyl-PEG24-azide (mPEG azide where m=24 in reaction formula (3)), DMSO (SP grade), and 20 mM CuSO ４ A solution of 2 mg / mL of CEL-C≡CH, 2 mM of methyl-PEG24-azide, 2 mM of CuSO 4 , and 8 mM of sodium ascorbate in ultrapure water was used. ４ The mixture was added to a 5 mL tube so that the concentration of DMSO was 0.5 mM and that of ascorbic acid was 0.2 mM, and the mixture was stirred at 25°C for 1 hour to react. The volume of the reaction solution was 5000 µL, with the water / DMSO (volume ratio) being 1 / 1. After the reaction, the liquid was centrifuged (20,400 x g, 5 minutes, 4°C), the supernatant was removed, and then ultrapure water was added to redisperse the product. This centrifugal separation (under the same conditions) was repeated five times to obtain the cellulose oligomer aggregate (mEG) of Synthesis Example 1. ２４ _1)was obtained.

[0081] Synthesis Example 2: 4 mM of methyl-PEG24-azide, CuSO４ The amounts of cellulose oligomer aggregate (mEG) of Synthesis Example 2 were set to 1.0 mM and ascorbic acid to 0.4 mM, and the other conditions were the same as in Synthesis Example 1. ２４ _2) was obtained.

[0082] Synthesis Example 3: 6 mM of methyl-PEG24-azide, CuSO ４ The amounts of cellulose oligomers added were set to 1.5 mM and ascorbic acid to 0.6 mM, and the other conditions were the same as in Synthesis Example 1, and the cellulose oligomer aggregate (mEG ２４ _3)was obtained.

[0083] Synthesis Example 4 A CEL-C≡CH aqueous dispersion, 40 mM methyl-PEG12-azide (mPEG azide where m=12 in reaction formula (3)), DMSO (SP grade), and 20 mM CuSO ４ A solution of 2 mg / mL of CEL-C≡CH, 2 mM of methyl-PEG12-azide, 2 mM of CuSO 4, 8 mM of sodium ascorbate, and ultrapure water was added. ４ The mixture was added to a 5 mL tube so that the concentration of DMSO was 0.5 mM and that of ascorbic acid was 0.2 mM, and the mixture was stirred at 25°C for 1 hour to react. The water / DMSO (volume ratio) was 1 / 1, and the volume of the reaction solution was 2500 μL. After the reaction, the liquid was centrifuged (20,400 × g, 5 minutes, 4°C), the supernatant was removed, and then ultrapure water was added to redisperse the product. This centrifugal separation (under the same conditions) was repeated five times to obtain the cellulose oligomer aggregate (mEG) of Synthesis Example 4. １２ _1)was obtained.

[0084] Cellulose oligomer aggregates (mEG ２４ _1-3 and mEG １２Mass spectra were obtained for sample 1) by MALDI-TOF-MS. The results are shown in Figure 4. In Figure 4, the peaks surrounded by dashed lines at DP = 6 to 11 are peaks of CEL-C≡CH to which no PEG chains have been introduced, and the numbers 6 to 11 represent the degree of polymerization of cellulose in CEL-C≡CH. The peaks surrounded by dashed lines at DP = 6' to 10' are peaks of a PEG-introduced cellulose oligomer to which a PEG chain with m = 24 has been introduced, and the numbers 6' to 10' represent the degree of polymerization of cellulose in the PEG-introduced cellulose oligomer. The peaks surrounded by dashed lines at DP = 7" to 9" are peaks of a PEG-introduced cellulose oligomer to which a PEG chain with m = 12 has been introduced, and the numbers 7" to 9" represent the degree of polymerization of cellulose in the PEG-introduced cellulose oligomer.

[0085] From the integral values ​​of the peaks of the PEG-introduced cellulose oligomer and the peaks of CEL-C≡CH, each cellulose oligomer assembly (mEG ２４ _1-3 and mEG １２ The PEG introduction rate of each of the cellulose oligomers (1) was determined. The results are shown in Table 1 below. The PEG introduction rate is the proportion (mol %) of PEG-introduced cellulose oligomers to all cellulose oligomers constituting the cellulose oligomer assembly.

[0086]

[0087] Cellulose oligomer aggregates (mEG ２４ _1-3 and mEG １２ ATR / FT-IR measurements were performed on cellulose nanosheet 1), and the spectrum shown in Figure 5 was obtained. The spectrum obtained showed peaks derived from the hydroxyl groups of the cellulose forming the cellulose type II crystalline structure, peaks derived from the acetylene groups, and peaks derived from the amorphous PEG chains. This indicates that the click reaction does not affect the cellulose crystalline structure, and that the PEG chains exposed on the surface of the cellulose nanosheet are in an amorphous state.

[0088] [Reagents] The reagents BSA and OPD used in the following test examples are as follows: BSA: bovine serum albumin, manufactured by Fujifilm Wako Pure Chemical Industries, Ltd., protease-free OPD: o-phenylenediamine dihydrochloride, manufactured by Nacalai Tesque, for water quality analysis

[0089] Test Example 1: Evaluation of binding of anti-PEG antibody (PEG-B-47) PEG-B-47 (manufactured by ABCAM) was used as the primary anti-PEG antibody. PEG-B-47 is a methoxy-terminated PEG-selective monoclonal anti-PEG antibody of rabbit species and IgG class, which specifically binds to PEG chains of 140 Da (DP=3) or more having a methoxy group at the end. PEG-B-47 was diluted with a 3.3% by mass BSA / PBS (pH 7.4) solution to prepare a primary antibody solution containing 110 ng / mL of PEG-B-47.

[0090] The cellulose oligomer aggregates (mEG) obtained in Synthesis Examples 1 and 4 ２４ _1 and mEG １２ Two types of cellulose dispersions according to the examples were prepared by dispersing each of the above-synthesized cellulose oligomer aggregates (CEL-C≡CH) having propargyl groups in PBS at a concentration of 5 mg / mL. A cellulose dispersion according to the comparative example was also prepared by dispersing the above-synthesized cellulose oligomer aggregate (CEL-C≡CH) in PBS at a concentration of 5 mg / mL.

[0091] The secondary antibody used to evaluate the binding of the anti-PEG antibody was an anti-rabbit IgG, HRP-labeled antibody (manufactured by GeneTex), which was diluted with a 3.3% by mass BSA / PBS solution to prepare a secondary antibody solution with a secondary antibody content of 440 ng / mL.

[0092] 1. Antigen-antibody reaction of primary antibody: 10 μL of cellulose dispersion was added to a 0.2 mL PCR tube, followed by 100 μL of primary antibody solution. After dispersion, the mixture was incubated at 25°C for 1 hour to carry out an antigen-antibody reaction. The primary antibody antigen-antibody reaction system had a cellulose oligomer assembly concentration of 0.45 mg / mL, a primary antibody concentration of 100 ng / mL, and a BSA concentration of 3% by mass. After incubation, the mixture was centrifuged (15,000 rpm, 25°C, 5 minutes), the supernatant was removed, and the mixture was redispersed in 100 μL of PBS. This procedure was repeated three times for washing.

[0093] 2. Antigen-antibody reaction of secondary antibodies After washing, i.e., after centrifugation and removal of the supernatant, 100 μL of secondary antibody solution was added to the PCR tube and dispersed, followed by incubation at 25°C for 1 hour to carry out an antigen-antibody reaction. The system for this secondary antibody antigen-antibody reaction had a cellulose oligomer assembly concentration of 0.45 mg / mL and a secondary antibody concentration of 400 ng / mL. After incubation, the tube was centrifuged (15,000 rpm, 25°C, 5 minutes), the supernatant was removed, and the tube was redispersed in 100 μL of PBS. This procedure was repeated three times for washing.

[0094] 3. Enzyme Reaction Next, after washing, i.e., after centrifugation and removal of the supernatant, 100 μL of substrate solution was added to the PCR tube, dispersed, and incubated at 25°C for 30 minutes. The substrate solution used was citrate phosphate buffer (pH 4.0) containing 2 mg / mL OPD and 0.11 v / v% hydrogen peroxide. After incubation, 100 μL of 3N sulfuric acid was added, and the mixture was centrifuged (15,000 rpm, 25°C, 10 minutes). 100 μL of the supernatant was added to a 96-well plate, and the absorbance at 490 nm was measured using a plate reader (BioTek "Synergy H1").

[0095] The results are shown in Figure 6. In Figure 6, the graph on the left, "PEG-B-47(+)", shows the "mEG ２４ _1" and "mEG １２ "_1" is the result of the above test. The test was carried out in three sets of three experiments, and the average absorbance of the average value of each set was calculated.

[0096] As a control, reactions 1 to 3 above were carried out in the same manner as above, except that 100 μL of a 3.3% by mass BSA / PBS (pH 7.4) solution containing no PEG-B-47 was added instead of 100 μL of the primary antibody solution in the antigen-antibody reaction of the primary antibody. The results are shown as the graph on the right side of Figure 6, "PEG-B-47(-)."

[0097] Furthermore, in the antigen-antibody reaction of the primary antibody, 10 μL of PBS containing no cellulose oligomer aggregate was added instead of 10 μL of the cellulose dispersion, and the other conditions were the same as in the reactions 1 to 3. The results are shown as "(-)" in the graphs for "PEG-B-47(+)" and "PEG-B-47(-)" in Figure 6.

[0098] As shown in Figure 6, the cellulose oligomer assembly CEL-C≡CH according to the comparative example had the same absorbance as the "(-)" cellulose oligomer-free assembly, and the anti-PEG antibody PEG-B-47 did not bind. ２４ _1 or mEG １２ In the case of (_1), the absorbance is significantly higher than that of "(-)", indicating that the anti-PEG antibody PEG-B-47 is bound. Therefore, in the case of the cellulose oligomer assembly of the example, the methoxy-terminated PEG-selective anti-PEG antibody can be detected.

[0099] Test Example 2: Evaluation of binding of anti-PEG antibody (15-2b) 15-2b (manufactured by Merck) was used as the anti-PEG antibody, which was the primary antibody. 15-2b is a methoxy-terminated PEG-selective monoclonal anti-PEG antibody of mouse species and IgG2b class, which specifically binds to PEG chains of 560 Da (DP = 12) or more having a methoxy group at the end. 15-2b was diluted with a 3.3% by mass BSA / PBS (pH 7.4) solution to prepare a primary antibody solution containing 15-2b at 110 ng / mL.

[0100] The cellulose oligomer aggregates (mEG) obtained in Synthesis Examples 1 to 4 ２４ _1-3 and mEG １２Four types of cellulose dispersions according to the examples were prepared by dispersing each of the above-synthesized cellulose oligomer aggregates (CEL-C≡CH) having propargyl groups in PBS at a concentration of 5 mg / mL. A cellulose dispersion according to the comparative example was also prepared by dispersing the above-synthesized cellulose oligomer aggregate (CEL-C≡CH) in PBS at a concentration of 5 mg / mL.

[0101] The secondary antibody used in evaluating the binding of the anti-PEG antibody was an anti-mouse IgG, HRP-labeled antibody (manufactured by Sigma-Aldrich), which was diluted with a 3.3% by mass BSA / PBS solution to prepare a secondary antibody solution with a secondary antibody content of 440 ng / mL.

[0102] 1. Antigen-antibody reaction of primary antibody: 10 μL of cellulose dispersion was added to a 0.2 mL PCR tube, followed by 100 μL of primary antibody solution. After dispersion, the mixture was incubated at 25°C for 1 hour to carry out an antigen-antibody reaction. The primary antibody antigen-antibody reaction system had a cellulose oligomer assembly concentration of 0.45 mg / mL, a primary antibody concentration of 100 ng / mL, and a BSA concentration of 3% by mass. After incubation, the mixture was centrifuged (15,000 rpm, 25°C, 5 minutes), the supernatant was removed, and the mixture was redispersed in 100 μL of PBS. This procedure was repeated three times for washing.

[0103] 2. Antigen-antibody reaction of secondary antibodies After washing, i.e., after centrifugation and removal of the supernatant, 100 μL of secondary antibody solution was added to the PCR tube and dispersed, followed by incubation at 25°C for 1 hour to carry out an antigen-antibody reaction. The system for this secondary antibody antigen-antibody reaction had a cellulose oligomer assembly concentration of 0.45 mg / mL and a secondary antibody concentration of 400 ng / mL. After incubation, the tube was centrifuged (15,000 rpm, 25°C, 5 minutes), the supernatant was removed, and the tube was redispersed in 100 μL of PBS. This procedure was repeated three times for washing.

[0104] 3. Enzyme Reaction Next, after washing, i.e., after centrifugation and removal of the supernatant, 100 μL of substrate solution was added to the PCR tube, dispersed, and incubated at 25°C for 30 minutes. The substrate solution used was citrate phosphate buffer (pH 4.0) containing 2 mg / mL OPD and 0.11 v / v% hydrogen peroxide. After incubation, 100 μL of 3N sulfuric acid was added, and the mixture was centrifuged (15,000 rpm, 25°C, 10 minutes). 100 μL of the supernatant was added to a 96-well plate, and the absorbance at 490 nm was measured using a plate reader (BioTek "Synergy H1").

[0105] The results are shown in Figure 7. In Figure 7, the graph on the left, "15-2b(+)" shows "mEG ２４ _1", "mEG" ２４ _2", "mEG" ２４ _3", "mEG" １２ The results of the above test are "CEL-C≡CH" and "CEL-C≡_1". The test was carried out in three sets of three experiments, and the average absorbance of each set was calculated.

[0106] As a control, reactions 1 to 3 above were carried out in the same manner as above, except that 100 μL of a 3.3% by mass BSA / PBS (pH 7.4) solution not containing 15-2b was added instead of 100 μL of the primary antibody solution in the antigen-antibody reaction of the primary antibody. The results are shown as the graph "15-2b(-)" on the right side of Figure 7.

[0107] Furthermore, in the antigen-antibody reaction of the primary antibody, 10 μL of PBS containing no cellulose oligomer aggregate was added instead of 10 μL of the cellulose dispersion, and the other conditions were the same as in the reactions 1 to 3. The results are shown as "(-)" in the graphs "15-2b(+)" and "15-2b(-)" in Figure 7.

[0108] As shown in Figure 7, the cellulose oligomer assembly CEL-C≡CH according to the comparative example had the same absorbance as the "(-)" cellulose oligomer-free assembly, and anti-PEG antibody 15-2b did not bind. In contrast, the cellulose oligomer assembly (mEG ２４ _1-3 or mEG １２In the case of (_1), the absorbance is significantly higher than that of "(-)", indicating that anti-PEG antibody 15-2b is bound. Therefore, in the case of the cellulose oligomer assembly of the example, the methoxy-terminated PEG-selective anti-PEG antibody can be detected.

[0109] Test Example 3: Evaluation of binding of anti-PEG antibody (6.3) 6.3 (manufactured by Merck) was used as the anti-PEG antibody, which is the primary antibody. 6.3 is a backbone-selective anti-PEG antibody of animal species: mouse, class: IgG1, which specifically binds to PEG chains of 750 Da (DP = 16) or more (Samuel K. Lai, Commun. Chem. 2020, 3, 124). 6.3 was diluted with a 3.3% by mass BSA / PBS (pH 7.4) solution to prepare a primary antibody solution containing 110 ng / mL of 6.3. The cellulose dispersion was prepared in the same manner as in Test Example 1 using the cellulose oligomer of Synthesis Example 1. The same secondary antibody solution as in Test Example 1 was used.

[0110] 1. Antigen-antibody reaction of primary antibody: 10 μL of cellulose dispersion was added to a 0.2 mL PCR tube, followed by 100 μL of primary antibody solution. After dispersion, the mixture was incubated at 25°C for 1 hour to carry out an antigen-antibody reaction. The primary antibody antigen-antibody reaction system had a cellulose oligomer assembly concentration of 0.45 mg / mL, a primary antibody concentration of 100 ng / mL, and a BSA concentration of 3% by mass. After incubation, the mixture was centrifuged (15,000 rpm, 25°C, 5 minutes), the supernatant was removed, and the mixture was redispersed in 100 μL of PBS. This procedure was repeated three times for washing.

[0111] 2. Antigen-antibody reaction of secondary antibodies After washing, i.e., after centrifugation and removal of the supernatant, 100 μL of secondary antibody solution was added to the PCR tube and dispersed, followed by incubation at 25°C for 1 hour to carry out an antigen-antibody reaction. The system for this secondary antibody antigen-antibody reaction had a cellulose oligomer assembly concentration of 0.45 mg / mL and a secondary antibody concentration of 400 ng / mL. After incubation, the tube was centrifuged (15,000 rpm, 25°C, 5 minutes), the supernatant was removed, and the tube was redispersed in 100 μL of PBS. This procedure was repeated three times for washing.

[0112] 3. Enzyme Reaction Next, after washing, i.e., after centrifugation and removal of the supernatant, 100 μL of substrate solution was added to the PCR tube, dispersed, and incubated at 25°C for 30 minutes. The substrate solution used was citrate phosphate buffer (pH 4.0) containing 2 mg / mL OPD and 0.11 v / v% hydrogen peroxide. After incubation, 100 μL of 3N sulfuric acid was added, and the mixture was centrifuged (15,000 rpm, 25°C, 10 minutes). 100 μL of the supernatant was added to a 96-well plate, and the absorbance at 490 nm was measured using a plate reader (BioTek "Synergy H1").

[0113] The results are shown in Figure 8. In Figure 8, the graph on the left, "6.3 (+)" indicates "mEG ２４ The results of the above test are "CEL-C≡CH" and "CEL-C≡_1". The test was carried out in three sets of three experiments, and the average absorbance of each set was calculated.

[0114] As a control, reactions 1 to 3 above were carried out in the same manner as above, except that 100 μL of a 3.3% by mass BSA / PBS (pH 7.4) solution not containing 6.3 was added instead of 100 μL of the primary antibody solution in the antigen-antibody reaction of the primary antibody. The results are shown as the graph "6.3(-)" on the right side of Figure 8.

[0115] Furthermore, in the antigen-antibody reaction of the primary antibody, 10 μL of PBS containing no cellulose oligomer aggregate was added instead of 10 μL of the cellulose dispersion, and the other conditions were the same as in the reactions 1 to 3. The results are shown as "(-)" in the graphs "6.3(+)" and "6.3(-)" in Figure 8.

[0116] As shown in Figure 8, the cellulose oligomer assembly CEL-C≡CH according to the comparative example had almost the same absorbance as the "(-)" cellulose oligomer-free assembly, and anti-PEG antibody 6.3 did not bind. In contrast, the cellulose oligomer assembly (mEG ２４ In the case of (_1), the absorbance is higher than that of "(-)", indicating that anti-PEG antibody 6.3 is bound. Therefore, in the case of the cellulose oligomer assembly of the example, backbone-selective anti-PEG antibodies can be detected.

[0117] The various numerical ranges described in this specification can be arbitrarily combined with their respective upper and lower limit values, and all such combinations are considered to be preferred numerical ranges described in this specification. Furthermore, a numerical range described as "X to Y" means from X to Y.

[0118] Although several embodiments of the present invention have been described above, these embodiments are presented as examples and are not intended to limit the scope of the invention. These embodiments can be implemented in various other forms, and various omissions, substitutions, and modifications can be made without departing from the spirit of the invention. These embodiments and their omissions, substitutions, modifications, etc. are included within the scope and spirit of the invention, as well as within the scope of the invention described in the claims and their equivalents.

Claims

1. General formula (1): In the formula (1), n ​​is an average degree of polymerization and is a number from 6 to 16, m is an average degree of polymerization and is a number not less than 3, and A is a divalent organic group connecting an oxygen atom bonded to the anomeric carbon at the reducing end of the cellulose oligomer to a polyethylene glycol chain, the divalent organic group having 1 to 24 carbon atoms and containing at least one heteroatom selected from the group consisting of nitrogen atoms, oxygen atoms, and sulfur atoms.

2. The anti-PEG antibody binding material according to claim 1, wherein A in formula (1) represents a divalent organic group having 2 to 10 carbon atoms containing at least one group selected from the group consisting of a triazole group, an amide group, and a thioether group.

3. A in the formula (1) is where R １ represents a hydrocarbon group having 1 to 10 carbon atoms, R ２ The anti-PEG antibody binding material of claim 1 , wherein represents a single bond or a hydrocarbon group having 1 to 10 carbon atoms.

4. The anti-PEG antibody binding material according to claim 1, wherein m in formula (1) is a number of 10 or more.

5. A biosensor comprising an anti-PEG antibody binding material according to any one of claims 1 to 4.

6. An adsorbent comprising the anti-PEG antibody binding material according to any one of claims 1 to 4.

7. General formula (1): a divalent organic group having 1 to 24 carbon atoms and containing at least one heteroatom selected from the group consisting of nitrogen atoms, oxygen atoms, and sulfur atoms, connecting an oxygen atom bonded to the anomeric carbon at the reducing end of the cellulose oligomer to a polyethylene glycol chain; incubating the cellulose oligomer after incubation with the specimen with a sample containing the secondary antibody under conditions that allow binding of the secondary antibody to the anti-PEG antibody; and detecting the anti-PEG antibody contained in the specimen by using the cellulose oligomer after incubation with the sample with the secondary antibody as a probe.

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