Method for stabilizing hemoglobin A2 and hemoglobin F, method for producing hemoglobin solution, and hemoglobin solution

Abstract

Provided are: a method for stabilizing hemoglobin A2 and hemoglobin F, whereby hemoglobin A2 and hemoglobin F in a hemoglobin solution can be stabilized; a method for producing a hemoglobin solution; and a hemoglobin solution. A method for stabilizing hemoglobin A2 and hemoglobin F, and a method for producing a hemoglobin solution, the method comprising a step for preparing a solution containing a buffer, sucrose, and hemoglobin, the pH of the solution being 6.0-9.6, the concentration of the buffer in the solution being 5-150 mM, and the concentration of the hemoglobin in the solution being 5-20 g / L, the hemoglobin at least comprises hemoglobin A2 and hemoglobin F; and a hemoglobin solution.

Description

Technical Field

[0001] The present invention relates to a method for stabilizing hemoglobin A2 and hemoglobin F, a method for producing a hemoglobin solution, and a hemoglobin solution. Background Art

[0002] At the scene of clinical examination, quality control substances are used to ensure the quality of the test results. Usually, quality control substances are sold in a freeze-dried state and dissolved in purified water or the like before use. The remaining quality control substances are refrigerated and stored for use later. Therefore, the quality control substances in liquid state are required to be stable for a long time.

[0003] Patent Document 1 describes a managed sample of hemoglobin (hereinafter also referred to as "Hb") F, which is characterized in that a hemolysate of human erythrocytes and a mixed hemolysate of human umbilical cord blood contain sucrose as a stabilizer.

[0004] Prior art literature

[0005] Patent Literature

[0006] Patent Document 1: Japanese Patent Application Publication No. 2016-075504 Summary of the invention

[0007] Problems to be solved by the invention

[0008] Patent Document 1 describes that the charge of Hb molecules varies depending on the type, so there are Hb stabilized by sucrose and Hb that is unstable. Therefore, it is considered difficult to simultaneously stabilize different types of Hb by simply adding sucrose.

[0009] The present invention relates to providing a method for stabilizing HbA2 and HbF, a method for producing an Hb solution, and an Hb solution, which are capable of stabilizing HbA2 and HbF in an Hb solution.

[0010] Methods used to solve problems

[0011] Methods for solving the above-mentioned problems include the following.

[0012] <1> A method for stabilizing hemoglobin A2 and hemoglobin F, comprising the steps of preparing a solution containing a buffer, sucrose and hemoglobin,

[0013] The pH of the above solution is 6.0 to 9.6.

[0014] The concentration of the buffer in the solution is 5 to 150 mM.

[0015] The concentration of the hemoglobin in the solution is 5 to 20 g / L.

[0016] The above-mentioned hemoglobin includes at least hemoglobin A2 and hemoglobin F.

[0017] <2> The method for stabilizing hemoglobin A2 and hemoglobin F according to <1>, wherein the concentration of the sucrose in the solution is 5 to 20% (w / v).

[0018] <3> A method for producing a hemoglobin solution, comprising the steps of preparing a solution containing a buffer, sucrose and hemoglobin,

[0019] The pH of the above solution is 6.0 to 9.6.

[0020] The concentration of the buffer in the solution is 5 to 150 mM.

[0021] The concentration of the hemoglobin in the solution is 5 to 20 g / L.

[0022] The above-mentioned hemoglobin includes at least hemoglobin A2 and hemoglobin F.

[0023] <4> The method for producing a hemoglobin solution according to <3>, wherein the concentration of the sucrose in the solution is 5 to 20% (w / v).

[0024] <5> A hemoglobin solution comprising a buffer, sucrose and hemoglobin, wherein:

[0025] The pH of the above solution is 6.0 to 9.6.

[0026] The concentration of the buffer in the solution is 5 to 150 mM.

[0027] The concentration of the hemoglobin in the solution is 5 to 20 g / L.

[0028] The above-mentioned hemoglobin includes at least hemoglobin A2 and hemoglobin F.

[0029] <6> The hemoglobin solution according to <5>, wherein the concentration of the sucrose in the solution is 5 to 20% (w / v).

[0030] <7> The hemoglobin solution according to <5> or <6>, which is used as a quality control substance.

[0031] Effects of the Invention

[0032] According to the present invention, there are provided a method for stabilizing HbA2 and HbF, a method for producing an Hb solution, and an Hb solution, which are capable of stabilizing HbA2 and HbF in an Hb solution. BRIEF DESCRIPTION OF THE DRAWINGS

[0033] Figure 1The chromatogram of high performance liquid chromatography performed in Example 1 under the conditions of 100 mM Tris-HCl (pH 8.2) and an Hb concentration of 10 g / L is shown.

[0034] Figure 2 The chromatogram of the high performance liquid chromatography performed in Example 1 under the conditions of 250 mM phosphate buffer (pH 6.0) and an Hb concentration of 5 g / L is shown. DETAILED DESCRIPTION

[0035] Below, the mode for implementing the embodiment of the present invention is described in detail. However, the embodiment of the present invention is not limited to the following embodiment. In the following embodiment, unless otherwise specified, its constituent elements (also including element steps, etc.) are not necessary. It is also the same for numerical values ​​and their ranges, and does not limit the embodiment of the present invention.

[0036] In the present invention, in a numerical range expressed using "to", the numerical values ​​described before and after "to" are included as the minimum value and the maximum value, respectively.

[0037] In the numerical range recorded in stages in the present invention, the upper limit or lower limit recorded in a numerical range can be replaced by the upper limit or lower limit of the numerical range recorded in other stages. In addition, in the numerical range recorded in the present invention, the upper limit or lower limit of this numerical range can also be replaced by the value shown in the embodiment.

[0038] In the present invention, a combination of two or more preferred embodiments is a more preferred embodiment.

[0039] In the present invention, the total Hb content in the solution is measured by the cyanomethemoglobin method. Specifically, for example, the total Hb content can be measured using Hemoglobin Test Wako (Fujifilm Wako Pure Chemical Industries, Ltd.).

[0040] In the present invention, the ratio of HbA2 to HbF in Hb is measured by peak area ratio in high performance liquid chromatography.

[0041] In the description of the embodiments of the present invention, pH refers to pH at 25° C. unless otherwise specified.

[0042] <Method for stabilizing HbA2 and HbF>

[0043] The method for stabilizing HbA2 and HbF of the present invention comprises the step of preparing a solution containing a buffer, sucrose and Hb, wherein the pH of the solution is 6.0 to 9.6, the concentration of the buffer in the solution is 5 to 150 mM, the concentration of the Hb in the solution is 5 to 20 g / L, and the Hb contains at least HbA2 and HbF. In the present invention, the prepared solution containing a buffer, sucrose and Hb is also described as "Hb solution".

[0044] The inventors have found that by preparing Hb solution under the above-mentioned specific conditions, even if Hb solution is stored for a long time, both HbA2 and HbF can be stabilized. The charge of Hb varies according to the type, and it is speculated that its stabilization conditions are also different, but in the solution where pH, buffer concentration and Hb concentration are within the above range and containing sucrose, the charge state of HbA2 and HbF is difficult to change, so it is speculated that both can exist stably for a long time. Thus, for example, a precision management material containing long-term stable HbA2 and HbF can be manufactured. In addition, for example, refrigerated storage can be performed from the preparation (dissolution) of the precision management material to the end of use. Therefore, the cost of precision management can be reduced when precision management is implemented daily, and steps such as preparation during use and subdivided frozen storage after preparation can be omitted, so the handling property is also improved. However, the embodiments of the present invention are not subject to any limitation of the above-mentioned mechanism of action. In the present invention, HbA2 and HbF "stable" means that before and after the storage of the Hb solution, the change of the ratio (%) of HbA2 and HbF relative to total Hb is small, that is, within a predetermined range.

[0045] In patent document 1, it is recorded that although sucrose shows a stabilizing effect on HbA1c and HbF, no stabilizing effect brought by sucrose that can withstand practicality is found for HbA1A. In addition, it is recorded that the charge of Hb is different according to the type, the difference in charge has a great impact on the detection state of high performance liquid chromatography, and the stabilizer of Hb is not easy to affect the state of Hb charge, which becomes a condition, but it is difficult to predict according to the correlation between the state of Hb charge and the carbohydrate molecule used as a stabilizer. On the other hand, the inventor focuses on HbA2 and HbF, two types of Hb, and studies the method for making both long-term stabilization. According to the method of the present invention, it is particularly useful to make HbA2 and HbF all long-term stabilization.

[0046] (Hemoglobin)

[0047] The Hb solution contains at least HbA2 and HbF, and may or may not contain other Hb such as HbA0 and HbA1. Typically, the Hb solution contains other Hb such as HbA0 and HbA1 derived from blood.

[0048] The concentration of Hb in the Hb solution (i.e., total Hb concentration) is 5 to 20 g / L. From the viewpoint of stabilizing HbA2 and HbF better, the Hb concentration in the Hb solution is preferably 10 to 20 g / L, and more preferably 10 to 18 g / L. Among them, when the buffer described below is a phosphate buffer, the Hb concentration is preferably 10 to 20 g / L, and more preferably 10 to 18 g / L. When the buffer is a Tris buffer, the Hb concentration is preferably 10 to 20 g / L. When the buffer is a carbonate-bicarbonate buffer, the Hb concentration is preferably 10 to 20 g / L.

[0049] The sources of HbA2 and HbF are not particularly limited, and examples thereof include human and non-human animal blood, preferably human blood. Examples of non-human animals include non-human mammals (monkeys, dogs, cats, mice, rats, rabbits, cows, horses, pigs, sheep, etc.), birds (chickens, quails, etc.), etc.

[0050] HbA2 may be, for example, HbA2 obtained from human peripheral blood. HbF may be, for example, HbF obtained from human peripheral blood or human umbilical cord blood rich in HbF. The sources of HbA2 and HbF may be the same sample or different samples. It is simple to prepare an Hb solution containing HbA2 and HbF from the same sample. In addition, by mixing HbA2 and HbF obtained from different samples, it is easy to arbitrarily adjust the concentration ratio of HbA2 and HbF.

[0051] For example, HbA2 and HbF can be obtained by hemolyzing the blood of the above-mentioned human or non-human animal and extracting Hb. Hemolysis can be performed, for example, by adding EDTA to the precipitate obtained by centrifuging the whole blood obtained by blood collection, and then freezing and thawing. In addition, HbA2 and HbF can also be obtained by purifying the solution obtained by hemolysis as described above by dialysis or the like.

[0052] (Buffer)

[0053] The Hb solution contains a buffer. Examples of the buffer include phosphate buffer, Tris buffer (e.g., Tris-HCl, TBS, TBS(-)-T, TAE, TBE, TE, etc.), carbonate-bicarbonate buffer, etc. From the perspective of better stabilization of HbA2 and HbF, Tris buffer is preferred, and Tris-HCl is more preferred.

[0054] The concentration of the buffer in the Hb solution is 5 to 150 mM, preferably 5 to 100 mM, more preferably 5 to 50 mM, and further preferably 5 to 30 mM from the viewpoint of better stabilization of HbA2 and HbF. Among them, when the buffer is a phosphate buffer, the concentration of the buffer is preferably 5 to 100 mM, more preferably 5 to 50 mM. When the buffer is a Tris buffer, the concentration of the buffer is preferably 5 to 100 mM. When the buffer is a carbonate-bicarbonate buffer, the concentration of the buffer is preferably 5 to 100 mM, more preferably 5 to 50 mM, and further preferably 5 to 30 mM.

[0055] (sucrose)

[0056] The Hb solution contains sucrose. Sucrose functions as a stabilizer for Hb. The concentration of sucrose in the solution is preferably 5 to 20% (w / v), more preferably 6 to 18% (w / v), and even more preferably 7 to 15% (w / v).

[0057] The amount of sucrose is preferably 5 to 20 times, more preferably 6 to 18 times, and even more preferably 7 to 15 times, the amount of Hb in the Hb solution in terms of mass ratio.

[0058] (Hb solution)

[0059] The pH of the Hb solution is 6.0 to 9.6, preferably 7.0 to 9.2, and more preferably 7.5 to 9.0 from the viewpoint of stabilizing HbA2 and HbF more favorably.

[0060] From the viewpoint of better stabilizing HbA2 and HbF, the pH of the Hb solution is 7.0 to 9.2 (more preferably 7.0 to 9.0), the concentration of the buffer is 5 to 150 mM (preferably 5 to 100 mM, more preferably 5 to 50 mM), and the Hb concentration is preferably 10 to 20 g / L (preferably 10 to 18 g / L).

[0061] The stabilization method of HbA2 and HbF of the present embodiment includes the step of preparing a solution containing a buffer, sucrose and Hb. The preparation method is not particularly limited, and the buffer, sucrose and Hb can be mixed by any method. In one embodiment, the Hb solution can be prepared by mixing a buffer, a sucrose solution, and a solution containing Hb containing HbA2 and HbF. In another embodiment, the Hb solution can be prepared by mixing a buffer, a sucrose solution, a solution containing HbA2 and a solution containing HbF. The order of mixing is not particularly limited.

[0062] For the prepared Hb solution, for example, when stored at 8°C after preparation, it is preferably stable for more than 21 days. For example, in the Hb solution, the change value of the ratio of HbA2 after 21 days of storage at 8°C after preparation is preferably less than 0.25%, more preferably less than 0.21%. In addition, in the Hb solution, the change value of the ratio of HbF after 21 days of storage at 8°C after preparation is preferably less than 0.60%, more preferably less than 0.40%, and further preferably less than 0.36%. The "change value" of the ratio of HbA2 or the ratio of HbF is obtained as follows. The ratio (%) of HbA2 and HbF in the Hb solution just after preparation and after storage is obtained by high performance liquid chromatography. The absolute difference between the above ratio after storage and the above ratio before storage is obtained, and each is used as the change value of the ratio (%) of HbA2 and HbF. In the determination of the change value, the conditions of high performance liquid chromatography are the method described in Example 1 or a method equivalent thereto.

[0063] <Method for producing hemoglobin solution>

[0064] The method for producing an Hb solution in one embodiment of the present invention includes the step of preparing a solution containing a buffer, sucrose and Hb, wherein the pH of the solution is 6.0 to 9.6, the concentration of the buffer in the solution is 5 to 150 mM, the concentration of the Hb in the solution is 5 to 20 g / L, and the Hb contains at least HbA2 and HbF. The details of the buffer, sucrose, Hb, Hb solution and the method for producing the same in this embodiment can all be applied to the matters described in the item "Method for stabilizing HbA2 and HbF".

[0065] <Hemoglobin solution>

[0066] The Hb solution in one embodiment of the present invention is an Hb solution containing a buffer, sucrose and Hb, wherein the pH of the solution is 6.0 to 9.6, the concentration of the buffer in the solution is 5 to 150 mM, the concentration of the Hb in the solution is 5 to 20 g / L, and the Hb contains at least HbA2 and HbF. The details of the buffer, sucrose, Hb, Hb solution and the preparation method thereof in this embodiment can all be applied to the matters described in the item "Method for stabilizing HbA2 and HbF".

[0067] In one embodiment, the Hb solution of this embodiment is used as a quality control substance. The quality control substance refers to a sample with a known composition for quality control of analysis, and the quality control substance of this embodiment refers to a sample with a known composition of HbA2 and HbF (i.e., the respective ratios (%) relative to the total Hb in the quality control substance).

[0068] [Example]

[0069] Next, the embodiment of the present invention will be specifically described by way of examples, but the embodiment of the present invention is not limited to these examples.

[0070] 1. Preparation of Hb solution

[0071] <Hemolysis>

[0072] (1) Human whole blood was centrifuged (10,000 g, 10 min, 4° C.) and the supernatant was discarded.

[0073] (2) Resuspend the precipitate in an equal volume of saline to that of whole blood.

[0074] (3) The suspension from step (2) was centrifuged (10,000 g, 10 min, 4° C.) and the supernatant was discarded.

[0075] (4) Perform steps (2) and (3) again.

[0076] (5) The precipitate was dissolved using 20 mM EDTA (pH 7.0) in a volume 1.3 times that of whole blood.

[0077] (6) Freeze the dissolved solution in step (5) at -80°C.

[0078] <Purification>

[0079] (7) After confirming that the dissolved solution in step (6) is completely frozen, thaw it in a water bath at room temperature (about 25°C) to prepare a hemolyzed sample.

[0080] (8) The hemolyzed sample was sealed in a dialysis membrane with a molecular weight cutoff of 3500 and dialyzed using purified water as the external solution.

[0081] (9) Replace the external solution (volume of external solution: more than 85 times of internal solution, once every 3 hours or more, for a total of 6 times).

[0082] (10) The recovered internal solution was centrifuged (40,000 g, 10 min, 4° C.) and the supernatant was recovered as solution A.

[0083] (11) The Hb concentration in solution A was measured using Hemoglobin Test Wako (Fujifilm Wako Pure Chemical Industries, Ltd.) (cyanomethemoglobin method).

[0084] (12) A 28% (w / v) sucrose solution, 0.5 M of each buffer solution, and the solution A determined in step (11) were mixed so that the final concentrations of the solutions became the specified concentrations.

[0085] 2. Evaluation

[0086] [Example 1]

[0087] <Test conditions>

[0088] ･Phosphate buffer: 5-250mM (pH 6.0, 7.0, 7.5, 8.0, 8.2)

[0089] ･Tris-HCl: 5~250mM (pH7.5, 8.0, 8.2, 9.0)

[0090] ･Carbonate-bicarbonate buffer: 5~250mM (pH9.2, 9.6)

[0091] ･Final sucrose concentration: 7% (w / v)

[0092] ･Hb concentration: 5-20 g / L

[0093] <High Performance Liquid Chromatography>

[0094] (1) The ratio of HbA2 to HbF in each sample was measured using a calibrated high performance liquid chromatography apparatus (n=3).

[0095] (2) The samples were stored at 8°C for 21 days.

[0096] (3) The ratio of HbA2 to HbF in each sample after storage was measured using a calibrated high performance liquid chromatography apparatus (n=3).

[0097] (4) The change values ​​before and after storage of the mean values ​​(n=3) of the measured values ​​of the HbA2 ratio (%) and the HbF ratio (%) were calculated.

[0098] <High Performance Liquid Chromatography Equipment>

[0099] (1) High performance liquid chromatography device

[0100] A commercially available cation exchange column filled with a hydrophilic polymer containing a methacrylate copolymer was connected to a commercially available high performance liquid chromatography apparatus, and an optical detector (specifically, an absorbance meter) for detecting the concentration of Hb flowing through the flow channel was installed at a predetermined position of the flow channel downstream of the cation exchange column.

[0101] (2) Eluent

[0102] Three types of eluents were prepared as aqueous solutions with the compositions shown in Table 1 below.

[0103] [Table 1]

[0104]

[0105] It should be noted that the pH of eluent A was adjusted to 5.08, the pH of eluent B was adjusted to 8.0, and the pH of eluent C was adjusted to 6.82. Regarding the elution power of Hb of these eluents, eluent A was the smallest, and eluent B was the largest.

[0106] (3) Chromatogram

[0107] The eluent A is passed through the high performance liquid chromatography apparatus of (1) above to equilibrate the column. Then, the hemolyzed blood sample is introduced into the column. Then, the eluent A is passed through for 13 seconds to elute HbA1a, HbA1b, HbF, and HbA1c. Next, a liquid obtained by mixing the eluent A and the eluent C at a ratio of 1:9 is passed through for 5 seconds to elute HbA0. Next, the eluent C is passed through for 5 seconds to elute HbA2. Then, the eluent B is passed through for 2 seconds to elute all the Hb remaining in the column, and then the eluent A is passed through for 5 seconds. A chromatogram is prepared based on the absorbance obtained by the optical detector at a detection wavelength of 420 nm during this elution.

[0108] The ratio (%) of HbA2 and the ratio (%) of HbF were calculated using the following formula.

[0109] HbA2 (%) = total area of ​​HbA2 / total area of ​​total Hb × 100

[0110] HbF (%) = total area of ​​HbF / total area of ​​total Hb × 100

[0111] <Calculation of change value>

[0112] The ratio (%) of HbA2 was calculated before and after the sample was stored at 8°C for 21 days. The absolute difference between the average value (n=3) after storage and the average value (n=3) before storage was calculated and used as the change value of the ratio (%) of HbA2. The change value of the ratio (%) of HbF was also calculated by the same method.

[0113] <Results>

[0114] ･The fluctuation values ​​of the ratio (%) of HbA2 and HbF are shown in the table below. In the table, "×" means that the measurement was performed, but the peak was outside the measurement range and no detection result was found. "-" means that the measurement was not performed. The fluctuation value of the ratio (%) of HbA2 is based on a range of less than 0.25%. The fluctuation value of the ratio (%) of HbF is based on a range of less than 0.60%.

[0115] [Table 2]

[0116]

[0117] [Table 3]

[0118]

[0119] [Table 4]

[0120]

[0121] [Table 5]

[0122]

[0123] [Table 6]

[0124]

[0125] [Table 7]

[0126]

[0127] An example of a chromatogram is shown in Figure 1 middle. Figure 1 The chromatogram is a chromatogram obtained under the conditions of 100 mM Tris-HCl (pH 8.2) and Hb concentration of 10 g / L. The dotted line represents the chromatogram on day 0 (preparation day), and the solid line represents the chromatogram on day 21. The peak of HbA2 and the amount of HbF are consistent on days 0 and 21, indicating that the state of the sample is stable.

[0128] Another example of a chromatogram is shown in Figure 2 middle. Figure 2 The chromatogram is a chromatogram obtained under the conditions of 250 mM phosphate buffer (pH 6.0) and Hb concentration of 5 g / L. The dotted line represents the chromatogram on day 0 (preparation day), and the solid line represents the chromatogram on day 21. The peaks of HbA2 do not match on day 0 and day 21, indicating that the state of the sample is unstable.

[0129] It can be seen from this example that in an Hb solution with a pH of 6.0 to 9.6, a buffer concentration of 5 to 150 mM, and an Hb concentration of 5 to 20 g / L, HbA2 and HbF are stable at 8°C for 21 days.

[0130] [Example 2]

[0131] <Test conditions>

[0132] ･Tris buffer: 25mM (pH8.8)

[0133] ･Final sucrose concentration: 7% (w / v)

[0134] ･Hb concentration: 13-18 g / L

[0135] <High Performance Liquid Chromatography>

[0136] (1) The ratio of HbA2 to HbF in each sample was measured using a calibrated high performance liquid chromatography apparatus (n=3).

[0137] (2) The samples were stored at 8°C for 21 days.

[0138] (3) The ratio of HbA2 to HbF in each sample after storage was measured using a calibrated high performance liquid chromatography apparatus (n=3).

[0139] (4) The change values ​​before and after storage of the mean values ​​(n=3) of the measured values ​​of the HbA2 ratio (%) and the HbF ratio (%) were calculated.

[0140] <High Performance Liquid Chromatography Equipment>

[0141] As a high performance liquid chromatography apparatus, an ADAMS HA-8180T (manufactured by ARKRAY, Inc.) was used. As an apparatus, a column Column Unit 80T (manufactured by ARKRAY, Inc.) filled with 0.45 ml of a hydrophilic polymer containing a methacrylate copolymer as a filler was used. As reagents, 80A was used as an eluent A for eluting Hb from the column, 80B was used as an eluent B, 80CT (manufactured by ARKRAY, Inc.) was used as an eluent C, and 80H was used as a hemolysis and washing liquid. The composition and pH of 80A, 80B, 80CT, and 80H are as follows. These reagents flowed into the column at a flow rate of 1.7 ml / min.

[0142] Eluent A (80A): Phosphate buffer (pH 5.35 ± 0.05)

[0143] Elution buffer B (80B): phosphate buffer (pH 8.05 ± 0.20)

[0144] Eluent C (80CT): Phosphate buffer (pH 7.0-7.4)

[0145] Hemolysis and washing solution (80H): Phosphate buffer containing surfactant (pH 7.5 ± 0.1)

[0146] <Calculation of change value>

[0147] In the same manner as in Example 1, the change value of the ratio (%) of HbA2 and HbF was determined.

[0148] <Results>

[0149] The variation values ​​of the ratio (%) of HbA2 and the ratio (%) of HbF of the sample relative to the initial value are shown in the table below. The variation value of the ratio (%) of HbA2 is less than 0.25% as the reference range. The variation value of the ratio (%) of HbF is less than 0.60% as the reference range. It was confirmed that the variation values ​​of HbA2 and HbF were both within the reference range.

[0150] [Table 8]

[0151]

Claims

1. A method for stabilizing hemoglobin A2 and hemoglobin F, comprising the steps of preparing a solution containing a buffer, sucrose and hemoglobin, The pH of the solution is 6.0 to 9.6, The concentration of the buffer in the solution is 5 to 150 mM, The concentration of the hemoglobin in the solution is 5 to 20 g / L, The hemoglobin includes at least hemoglobin A2 and hemoglobin F.

2. The method for stabilizing hemoglobin A2 and hemoglobin F according to claim 1, wherein: The concentration of the sucrose in the solution is 5-20 w / v %.

3. A method for producing a hemoglobin solution, comprising the steps of preparing a solution containing a buffer, sucrose and hemoglobin, The pH of the solution is 6.0 to 9.6, The concentration of the buffer in the solution is 5 to 150 mM, The concentration of the hemoglobin in the solution is 5 to 20 g / L, The hemoglobin includes at least hemoglobin A2 and hemoglobin F.

4. The method for producing a hemoglobin solution according to claim 3, wherein: The concentration of the sucrose in the solution is 5-20 w / v %.

5. A hemoglobin solution, which is a hemoglobin solution containing a buffer, sucrose and hemoglobin, wherein: The pH of the solution is 6.0 to 9.6, The concentration of the buffer in the solution is 5 to 150 mM, The concentration of the hemoglobin in the solution is 5 to 20 g / L, The hemoglobin includes at least hemoglobin A2 and hemoglobin F.

6. The hemoglobin solution according to claim 5, wherein The concentration of the sucrose in the solution is 5-20 w / v %.

7. The hemoglobin solution according to claim 5 or 6, which is used as a quality control substance.

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