Anti-CRP antibody-containing reagent for measuring blood coagulation time

Abstract

Provided is an anti-CRP antibody-containing specimen diluent for measuring blood coagulation time. Also provided is a reagent for measuring blood coagulation time that contains the specimen diluent. Also provided is a method for measuring blood coagulation time using the specimen diluent. Also provided is a method for evaluating the influence on blood coagulation time of CRP in a specimen using the specimen diluent.

Description

Reagent for measuring blood coagulation time containing an anti-CRP antibody

[0001] The present invention relates to a reagent for measuring blood coagulation time.

[0002] A blood coagulation test is a test for diagnosing a patient's blood coagulation ability by adding a reagent to a blood sample collected from the patient and measuring the blood coagulation time and the like. Typical examples of blood coagulation time include prothrombin time (PT) and activated partial thromboplastin time (APTT). When prolongation of the coagulation time of a blood sample is observed in a blood coagulation test, it is suspected to be caused by a decrease in coagulation factor activity, or a blood coagulation abnormality such as hemophilia or antiphospholipid antibody syndrome, and additional tests such as a cross-mixing test, coagulation factor activity quantification, and lupus anticoagulant quantification are performed for differentiation. Measurement of coagulation time is important as a primary screening test for detecting blood coagulation abnormalities in patients.

[0003] The coagulation time measured in a blood coagulation test can be affected by the properties of the blood sample and preparation procedures, such as contamination by tissue factor during blood collection, centrifugation conditions, and the presence of anticoagulants. In recent years, it has been reported that blood CRP (C-reactive protein) affects APTT, thereby impairing the accuracy of evaluation of a patient's blood coagulation ability (Non-Patent Documents 1 to 3). If accurate evaluation of blood coagulation ability cannot be performed, not only the patient's pathological condition cannot be detected, but there is also a risk of administering unnecessary treatment to the patient.

[0004] J Clin Lab Anal, 2022;36(9):e24608Clin Chem Lab Med, 2015, 53(5);e141-145J Clin Lab Anal, 2018;32(8);e22571

[0005] For accurate evaluation of blood coagulation ability, it is desired to reduce the influence of blood CRP on coagulation time in blood coagulation tests.

[0006] The present invention provides the following as representative embodiments: [1] A sample diluent for measuring blood coagulation time, containing an anti-CRP antibody. [2] The sample diluent according to [1], wherein the sample is plasma. [3] The sample diluent according to [1] or [2], wherein the anti-CRP antibody content is 250 to 20,000 μg / mL. [4] The sample diluent according to [1] or [2], wherein the sample diluent contains the anti-CRP antibody in an amount such that the concentration of the anti-CRP antibody in a mixture of the sample diluent and the sample is as follows. A = P × α A: Concentration of the anti-CRP antibody in a mixture of the sample diluent and the sample P: Concentration of CRP derived from the sample in a mixture of the sample diluent and the sample α: 1 to 6 [5] A sample diluent according to any one of [1] to [4], wherein the blood coagulation time is selected from the group consisting of prothrombin time, activated partial thromboplastin time, diluted prothrombin time, diluted partial thromboplastin time, kaolin coagulation time, and diluted Russell's snake venom time. [6] A lyophilized product of the sample diluent according to any one of [1] to [5]. [7] A reagent for measuring blood coagulation time, comprising the sample diluent according to any one of [1] to [5]. [8] A reagent for measuring blood coagulation time, comprising the lyophilized product according to [6]. [9] A method for measuring blood coagulation time, comprising measuring the blood coagulation time of a sample diluted with the sample diluent according to any one of [1] to [5]. A method for evaluating the effect of CRP in a sample on blood coagulation time, comprising: measuring the blood coagulation time Ta of a sample diluted with any one of the sample diluents described in item [1] to [5]; measuring the blood coagulation time Tb of the same sample diluted with a blood coagulation time measuring sample diluent that does not contain anti-CRP antibody, or without dilution; and comparing Ta and Tb.

[0007] This invention reduces the influence of CRP in a blood coagulation test on the coagulation time, thereby enabling accurate evaluation of the blood coagulation ability of the sample.

[0008] In this specification, "blood sample," "blood coagulation time," and "blood coagulation reaction" may be referred to simply as "sample," "coagulation time," and "coagulation reaction," respectively.

[0009] In coagulation time measurement, a sample solution containing a blood sample and a coagulation time measuring reagent is prepared, and the coagulation reaction of the sample solution is measured. The coagulation time measuring reagent generally consists of a first reagent containing an activator and a second reagent containing a coagulation initiator. The sample is incubated with the first reagent, and then the second reagent is added to initiate the coagulation reaction. The coagulation time of the sample solution after the addition of the second reagent is measured.

[0010] For measuring coagulation time, the subject's plasma is preferably used as the blood sample. Anticoagulants commonly used in coagulation tests may be added to the sample. For example, plasma can be obtained by collecting blood using a blood collection tube containing sodium citrate and then centrifuging it.

[0011] The present invention provides a sample diluent for measuring blood coagulation time. The sample diluent for measuring blood coagulation time of the present invention (hereinafter also referred to as "the diluent of the present invention") is used to dilute a sample to be used for measuring blood coagulation time. The sample mixed with the diluent of the present invention (hereinafter also referred to as the diluted sample) is subjected to coagulation time measurement after the first reagent and the second reagent are added as described above.

[0012] The diluent of the present invention contains an anti-CRP antibody. The anti-CRP antibody may be a monoclonal antibody, a polyclonal antibody, or a genetically engineered recombinant antibody, as long as it is an antibody that specifically binds to human CRP, and is preferably an anti-human CRP monoclonal antibody. The anti-CRP antibody can be prepared by conventional methods such as the hybridoma method. Alternatively, a commercially available anti-CRP antibody, such as a mouse anti-human CRP monoclonal antibody (e.g., catalog number: 66250-1-Ig; Proteintech Group, Inc.), can be used.

[0013] The content of the anti-CRP antibody in the diluent of the present invention is preferably 250 to 20,000 μg / mL, more preferably 500 to 10,000 μg / mL.

[0014] In another embodiment, the content of the anti-CRP antibody in the diluent of the present invention may be adjusted according to the CRP content in the sample. In a preferred embodiment, the diluent of the present invention contains the anti-CRP antibody in an amount such that the concentration of the anti-CRP antibody in the mixture of the diluent and the sample (i.e., the diluted sample) is as follows: A = P × α A: Concentration of the anti-CRP antibody in the mixture of the diluent of the present invention and the sample (μg / mL) P: Concentration of CRP derived from the sample in the mixture of the diluent of the present invention and the sample (μg / mL) α: 1 to 6, preferably 1 to 5, more preferably 1 to 4

[0015] The CRP concentration in a sample or diluted sample can be measured by conventional methods such as enzyme immunoassay (EIA), latex agglutination, or time-resolved fluorescence immunoassay (TRF). The concentration of anti-CRP antibody in the diluent or diluted sample can also be measured by conventional methods such as absorbance measurement or enzyme immunoassay (EIA).

[0016] The diluent of the present invention may further contain components that are commonly found in conventional sample diluents for measuring coagulation time, in addition to the anti-CRP antibody. Examples of such components include buffers, salts, and preservatives. As the buffer, a buffer having a buffering capacity in the pH range of 4 to 9, preferably 6 to 8, can be used. Examples of such buffers include Good's buffers such as PIPES, ACES, HEPES, TAPSO, POPSO, EPPS, and CHES, as well as amino acids, citric acid, phosphoric acid, acetic acid, imidazole, barbital, and GTA. Any one of these buffers can be used alone or in combination of two or more. The concentration of the buffer in the diluent of the present invention is not particularly limited as long as it can exhibit buffering capacity, but is preferably 1 to 200 mM, more preferably 10 to 100 mM. Examples of the salt include NaCl. The concentration of the salt in the diluent of the present invention is preferably 1 to 1000 mM, more preferably 10 to 200 mM. Examples of the aforementioned preservatives include ciprofloxacin, propionic acid, sodium benzoate, sodium azide, and a mixture of 2-methyl-1,2-thiazole-3(2H)-one and 5-chloro-2-methyl-1,2-thiazole-3(2H)-one (e.g., Proclin 300). These preservatives can be used individually or in combination of two or more. The concentration of the preservative in the diluted solution of the present invention is preferably 0.001 to 0.5 mM, more preferably 0.005 to 0.05 mM.

[0017] The diluent of the present invention may further contain additives for improving the storage or stability of the diluent or the anti-CRP antibody contained therein. Examples of such additives include proteins, sugars, amino acids, and surfactants.

[0018] The amount of the diluent of the present invention added to the sample is not particularly limited, but is preferably 0.1 to 15 μL, more preferably 0.5 to 5 μL, per 50 μL of sample.

[0019] The diluent of the present invention may be provided in liquid form, or in concentrated or solid form (e.g., lyophilized). The concentrations of the various components described above represent the concentrations in the diluent of the present invention in liquid form, ready for use.

[0020] In one embodiment, the diluent of the present invention is provided in the form of a liquid containing a predetermined amount of the anti-CRP antibody and can be used as is for diluting a sample. In one embodiment, the diluent may be provided as a refrigerated or frozen liquid. The frozen product is thawed before use. In another embodiment, the diluent of the present invention is provided in a concentrated or solidified form and can be diluted or restored before use to prepare the diluent of the present invention in the form of a liquid containing a predetermined amount of the anti-CRP antibody. For example, a lyophilized product of the diluent of the present invention may be provided and restored with purified water or the like for use as a sample diluent. In another embodiment, a reagent kit for preparing the diluent of the present invention may be provided, comprising the buffer and the anti-CRP antibody separately. By adding an appropriate amount of the anti-CRP antibody to the buffer, the diluent of the present invention containing a desired amount of anti-CRP antibody can be prepared. In yet another embodiment, a reagent kit for preparing the diluent of the present invention may be provided, comprising the diluent of the present invention, or its concentrated or solidified form, containing a predetermined amount of the anti-CRP antibody, and the anti-CRP antibody as an accessory. By adding an appropriate amount of the anti-CRP antibody to the diluent as needed, a diluent containing a desired amount of the anti-CRP antibody can be prepared according to the present invention.

[0021] The present invention also provides a reagent for measuring blood coagulation time, comprising the aforementioned diluent of the present invention. The reagent for measuring blood coagulation time of the present invention (hereinafter also referred to as "the reagent of the present invention") may contain the diluent of the present invention in a ready-to-use liquid form. Alternatively, the reagent of the present invention may include a concentrated solution of the diluent of the present invention, or a solidified product (e.g., lyophilized product) of the diluent of the present invention. Alternatively, the reagent of the present invention may include a reagent kit for preparing the diluent of the present invention, each containing the aforementioned buffer and anti-CRP antibody separately.

[0022] The reagent of the present invention may further contain various components necessary for measuring blood coagulation time. For example, the reagent of the present invention may contain various components necessary for the activation and coagulation of a sample. In one embodiment, the reagent of the present invention is a reagent kit comprising a first reagent containing an activator and / or a second reagent containing a coagulation initiator, which are used in the general coagulation time measurement reagents described above. In a preferred embodiment, the reagent of the present invention is a reagent kit comprising the first reagent and the second reagent.

[0023] For example, if the reagent of the present invention is a reagent for APTT measurement, the first reagent may contain a buffer, an activator, a phospholipid, etc. As the buffer, a buffer having a buffering capacity in the range of pH 4 to 9, preferably pH 6 to 8, can be used. Examples of the buffer include Good's buffers such as PIPES, ACES, HEPES, TAPSO, POPSO, EPPS, and CHES, as well as amino acids, citric acid, phosphoric acid, acetic acid, imidazole, barbital, and GTA. Any one of these buffers can be used alone or in combination of any two or more. The amount of the buffer in the first reagent is not particularly limited as long as it is an amount that exhibits buffering capacity, but is preferably 0.1 to 1000 mM, more preferably 0.5 to 500 mM, even more preferably 1 to 200 mM, and even more preferably 10 to 100 mM.

[0024] Examples of the activating agents include ellagic acid, kaolin, celite, colloidal silica, polyphenol compounds, anhydrous silicic acid, and metal ions. Examples of such metal ions include Zn. 2+ Mn 2+ ,Cd 2+ Fe 2+ Al 3+ These are some examples. The metal ion may be in the form of a salt. Any one of the activators listed above can be used alone or in combination of two or more. The content of the activator in the first reagent is not particularly limited, but is preferably 0.001 to 2 mg / mL, more preferably 0.001 to 0.5 mg / mL, even more preferably 0.001 to 0.1 mg / mL, and even more preferably 0.005 to 0.05 mg / mL.

[0025] Examples of the phospholipids mentioned above include natural or synthetic phospholipids. Examples of natural phospholipids include phospholipids derived from natural substances such as rabbit brain, bovine brain, human placenta, soybeans, and egg yolk. Examples of synthetic phospholipids include phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine (PS), phosphatidylglycerol (PG), phosphatidic acid (PA), phosphatidylinositol (PI), lysophosphatidylcholine (LPC), sphingomyelin (SM), and cardiolipin. Any one of these phospholipids can be used alone or in combination of two or more. For example, multiple phospholipids can be mixed and used as liposomes. The content of the phospholipid in the first reagent is not particularly limited, but is preferably 0.005 to 2 mM, more preferably 0.02 to 0.5 mM. In the APTT measurement reagent, the phospholipid, which is a platelet substitute, is added as a component necessary for the coagulation reaction. In the intrinsic coagulation pathway, phospholipids form complexes with activated factor IX and factor VIII, and these complexes activate the common factor system (factors V, II, and I) from factor X downwards, leading to fibrin formation.

[0026] The second reagent may contain a component that initiates blood coagulation, such as calcium ions. For example, the second reagent may contain water-soluble calcium compounds such as calcium chloride, calcium lactate, calcium gluconate, calcium glucuronate, and calcium tartrate. These calcium compounds can be used individually or in combination of two or more. The content of the calcium compound in the second reagent is not particularly limited, but is preferably 5 to 100 mM, more preferably 10 to 50 mM.

[0027] The first and second reagents may further contain additives to improve the shelf life or stability of the reagents. Examples of such additives include preservatives, antioxidants, dispersants, and stabilizers. Examples of preservatives include ciprofloxacin, propionic acid, sodium benzoate, sodium azide, and a mixture of 2-methyl-1,2-thiazole-3(2H)-one and 5-chloro-2-methyl-1,2-thiazole-3(2H)-one (e.g., Proclin 300). Examples of antioxidants include citric acid and butylhydroxyanisole. Examples of dispersants include phenol and collagen peptides. Examples of stabilizers include polyethylene glycol, polyvinylpyrrolidone, high molecular weight polysaccharides such as dextran and polysucrose (e.g., Ficol), salts such as sodium chloride, amino acids, and sugars.

[0028] The first and second reagents contained in the reagents of the present invention may be provided in liquid form, or in the form of concentrated or solidified material (e.g., lyophilized material). The concentrated or solidified material is dissolved in water, buffer solution, etc., at the time of use to prepare a liquid reagent. The concentrations of the various components mentioned above represent the concentrations in the reagent in liquid form that is ready for use.

[0029] A method for measuring blood coagulation time using the diluent or reagent of the present invention is provided. The method for measuring coagulation time according to the present invention (hereinafter also referred to as "the method of the present invention") includes measuring the blood coagulation time of a sample diluted with the diluent of the present invention. By using the diluent of the present invention, the method of the present invention reduces the influence of CRP in the sample on coagulation time, and thereby enables accurate evaluation of the blood coagulation ability of the sample.

[0030] Examples of coagulation times measured by the method of the present invention include prothrombin time (PT), activated partial thromboplastin time (APTT), diluted prothrombin time, diluted partial thromboplastin time, kaolin coagulation time, and diluted Russell's snake venom time. Of these, activated partial thromboplastin time (APTT) is preferred.

[0031] The method of the present invention can be carried out in accordance with conventional methods for measuring coagulation time, except that a sample diluted with the diluent of the present invention is used as the sample to be measured. For example, a preferred procedure for APTT measurement is as follows: The first reagent is mixed with plasma diluted with the diluent of the present invention, and the resulting mixture is heated. The heating conditions are, for example, 30°C to 40°C, preferably 35°C to 39°C. Then, the second reagent is added to the mixture to initiate the coagulation reaction. The coagulation reaction of the mixture (sample solution) after the addition of the second reagent is measured. For measurement, optical methods that measure the amount of scattered light, transmittance or absorbance of the sample solution, or mechanical methods that measure the viscosity of the sample solution may be used. The temperature of the sample solution during measurement is, for example, 30°C to 40°C, preferably 35°C to 39°C. The first and second reagents are preferably added to the sample at concentrations such that the clotting time (APTT) of normal plasma falls within the range of 15 to 80 seconds, preferably 15 to 60 seconds, and more preferably 20 to 50 seconds. The start time of the coagulation reaction can typically be defined as the time when the second reagent is mixed with the sample to initiate the coagulation reaction. Alternatively, other timings may be defined as the start time of the reaction. The duration for which the measurement of the coagulation reaction is continued is not particularly limited, but may be, for example, several tens of seconds to about 7 minutes from the time of mixing the sample and the second reagent. During this measurement time, the progress of the coagulation reaction can be repeatedly measured at predetermined intervals (e.g., photometry of scattered light intensity). For example, measurements may be taken at 0.1-second intervals. From the measured coagulation reaction, the clotting time (APTT) can be measured according to known methods.

[0032] The series of operations in the method of the present invention can be carried out using an automated analyzer. Alternatively, some of the operations may be performed manually. For example, a human may prepare the sample, and the subsequent operations can be carried out by an automated analyzer.

[0033] The specimens to which the method of the present invention is applied are not particularly limited, as long as they are specimens to which the risk of blood CRP affecting coagulation time is to be reduced.

[0034] The method of the present invention can be used to evaluate whether or not CRP in a sample affects the coagulation time. Therefore, the present invention further provides a method for evaluating the effect of CRP in a sample on coagulation time (hereinafter also referred to as "the evaluation method of the present invention"). This method includes measuring the coagulation time (Ta) of a sample diluted with the diluent of the present invention containing an anti-CRP antibody (diluent a), and measuring the coagulation time (Tb) of the same sample diluted with a diluent that does not contain an anti-CRP antibody (diluent b). Diluent b preferably has the same composition and pH as diluent a, except that it does not contain an anti-CRP antibody. Alternatively, the coagulation time of the same sample without dilution may be used as Tb. The measurement of coagulation time can be carried out according to a conventional method. The effect of CRP in the sample on coagulation time is evaluated by comparing Ta and Tb. More specifically, if Ta < Tb, it is suspected that CRP in the sample is prolonging the coagulation time. Patients with high blood CRP levels experience prolonged clotting time even if their coagulation ability is normal, which puts them at risk of being misdiagnosed with a coagulation disorder or receiving unnecessary treatment for it. By evaluating the effect of CRP in a sample on clotting time according to the present invention, it is possible to avoid the risk of misdiagnosis and unnecessary treatment for patients with high blood CRP levels as described above.

[0035] The specimens to which the evaluation method of the present invention is applied are not particularly limited, as long as they are specimens to which we want to investigate the possible influence of blood CRP on coagulation time. In one embodiment, specimens to which the evaluation method of the present invention is applied include specimens in which the coagulation time is near the border between the normal and abnormal ranges (e.g., APTT 40-45 seconds), the CRP value is high (e.g., ≥5 mg / dL), and there are no coagulation abnormality factors such as anticoagulant administration, factor VIII deficiency, or liver dysfunction, so the cause of the prolongation is unknown. Such specimens have or are suspected of having prolonged coagulation time, but it is difficult to determine whether the coagulation time near the border is due to a coagulation abnormality or to a false prolongation due to CRP, because it is unclear whether there are coagulation abnormality factors. In particular, when a patient with such a specimen whose cause of prolongation is unknown requires surgery, surgery may not be performed because the risks such as bleeding are unknown. The method of the present invention can clarify the bleeding risk of such specimens. Therefore, the specimens to which the evaluation method of the present invention is applied are preferably specimens from preoperative patients, and more preferably specimens from hospitalized preoperative patients with sufficient background information such as medication use. Furthermore, from the viewpoint of accuracy of the method of the present invention, the CRP value of the sample is preferably in the range of 5 to 20 mg / dL. Also, for samples with an abnormal coagulation time (for example, APTT of 45 seconds or more), other factors other than CRP, such as coagulation abnormalities, are suspected to be causing the prolongation, so it is desirable to undergo detailed examinations for coagulation abnormalities, etc., rather than confirming false prolongation by CRP using the method of the present invention.

[0036] For samples suspected of having a false prolongation of coagulation time due to CRP using the evaluation method of the present invention, additional tests can be performed to confirm whether or not factors other than CRP are influencing the coagulation time. Examples of such additional tests include cross-mixing tests, quantitative analysis of coagulation factor activity, quantitative analysis of lupus anticoagulant, and dRVVT measurement. For example, for samples suspected of having a false prolongation due to CRP, a cross-mixing test, quantitative analysis of coagulation factor activity, etc., can be performed in parallel with the method of the present invention, and the factors contributing to the coagulation time prolongation of the sample can be determined by comprehensively analyzing the results.

[0037] The present invention will be described in more detail below with reference to examples, but the present invention is not limited to these examples.

[0038] 1) Materials for preparing diluted samples: • Plasma: Warfarin-containing plasma (Access Bio LLC). • CRP: rhCRP solution (Oriental Yeast Co., Ltd., containing approximately 1 mg / mL of rhCRP in the CRP buffer). CRP buffer composition: 20 mmol / L Tris-HCl buffer (pH 7.5) containing 2 mmol / L CaCl2, 0.14 mol / L NaCl, and 0.05% NaN3. • Anti-CRP antibody: Mouse-derived anti-human monoclonal antibody (in-house prepared).

[0039] CRP-added samples were prepared by mixing a 3:1 (volume ratio) mixture of plasma and rhCRP solution with a 3:1 (volume ratio) mixture of plasma and CRP buffer in any desired ratio. A PBS solution containing anti-CRP antibody (antibody concentration 5550 μg / mL) was used as a diluent. Diluted samples were prepared by mixing a 9:1 (volume ratio) mixture of the CRP-added sample and the diluent with a 9:1 (volume ratio) mixture of the CRP-added sample and PBS in any desired ratio. The concentrations of CRP and anti-CRP antibody in the diluted samples are shown in Table 1.

[0040] 2) Reagents for APTT measurement: Reagent 1: Coagupia APTT-N APTT Reagent (Sekisui Medical Co., Ltd.) Reagent 2: Coagupia APTT-N Calcium Chloride Solution (Sekisui Medical Co., Ltd.)

[0041] 3) Measurement of coagulation time: 50 μL of diluted sample was heated in a cuvette at 37°C for 45 seconds, then 50 μL of the first reagent was added and heated for a further 171 seconds. 50 μL of the second reagent was added to the heated mixture to initiate the coagulation reaction. The reaction was carried out at 37°C. Light with a wavelength of 660 nm was irradiated onto the sample (sample solution) to which the second reagent had been added, and the amount of optical change due to the coagulation reaction (change in scattered light intensity) was measured. The coagulation time (APTT) was calculated from the measured coagulation reaction. The calculation of coagulation time was performed using an automated blood coagulation analyzer CP3000 (manufactured by Sekisui Medical Co., Ltd.).

[0042] Table 1 shows APTT measured from diluted specimens. APTT was prolonged as the CRP concentration in the diluted specimen increased, and it was confirmed that CRP prolongs APTT. APTT prolonged by CRP recovered as the concentration of anti-CRP antibody in the diluted specimen increased, and it was shown that the effect of CRP on APTT is mitigated by the anti-CRP antibody. It was observed that APTT is shortened when the antibody concentration is high. The antibody concentration in the diluted specimen required to correct APTT to within ±10% of the true value (43.8 seconds) (48.2 to 39.4 seconds) was [CRP concentration in diluted specimen × 1 to 6]. The antibody concentration in the diluted specimen required to correct APTT to within ±5% of the true value (43.8 seconds) (45.9 to 41.6 seconds) was [CRP concentration in diluted specimen × 1 to 4].

[0043]

Claims

1. A sample diluent for measuring blood coagulation time, containing anti-CRP antibody.

2. The sample diluent according to claim 1, wherein the sample is plasma.

3. The sample diluent according to claim 1, wherein the anti-CRP antibody content is 250 to 20,000 μg / mL.

4. The sample diluent according to claim 1, wherein the sample diluent contains the anti-CRP antibody in an amount such that the concentration of the anti-CRP antibody in the mixture of the sample diluent and the sample is as follows: A = P × α A: Concentration of the anti-CRP antibody in the mixture of the sample diluent and the sample P: Concentration of CRP derived from the sample in the mixture of the sample diluent and the sample α: 1 to 6 5. The sample diluent according to claim 1, wherein the blood coagulation time is selected from the group consisting of prothrombin time, activated partial thromboplastin time, diluted prothrombin time, diluted partial thromboplastin time, kaolin coagulation time, and diluted Russell's snake venom time.

6. A lyophilized product of a sample diluent according to any one of claims 1 to 5.

7. A reagent for measuring blood coagulation time, comprising a sample diluent according to any one of claims 1 to 5.

8. A reagent for measuring blood coagulation time, comprising the freeze-dried product described in claim 6.

9. A method for measuring blood coagulation time, comprising measuring the blood coagulation time of a sample diluted with the sample diluent described in any one of claims 1 to 5.

10. A method for evaluating the effect of CRP in a sample on blood coagulation time, comprising: measuring the blood coagulation time Ta of a sample diluted with the sample diluent according to any one of claims 1 to 5; measuring the blood coagulation time Tb of the sample diluted with a blood coagulation time measuring sample diluent that does not contain anti-CRP antibody, or without dilution; and comparing Ta and Tb.

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