Red blood cell or bone marrow cell iron release method, iron content detection method and kit

Abstract

The invention belongs to the field of determination of iron content in cells, and particularly relates to a red blood cell or bone marrow cell iron release method, an iron content detection method and a kit. The method comprises the following steps: treating red blood cells or bone marrow cells by using an aqueous solution containing a surfactant and nuclease to obtain a first treatment solution, digesting and removing DNA in the red blood cells or bone marrow cells by using the nuclease, and then adding an alkali solution into the first treatment solution for incubation to obtain a second treatment solution; a sodium hypochlorite solution is then added to the second treatment fluid to release iron in red blood cells or bone marrow cells. Compared with the prior art, the method has the advantages of being safe, rapid, accurate, easy to implement, low in cost and convenient to use and popularize.

Description

Technical Field

[0001] The invention belongs to the field of determination of iron content in cells, and particularly relates to a method for releasing iron from red blood cells or bone marrow cells, a method for detecting iron content and a kit. Background Art

[0002] Red blood cells are the most numerous blood cells in the blood and are responsible for transporting oxygen from the lungs to various tissues in the body. The number of red blood cells in adult males in my country is (4.0-5.5)×10 12 / L, for women: (3.5-5.0)×10 12 / L. The protein in red blood cells is mainly hemoglobin, which has four subunits, each of which has a heme, and each heme is bound to an iron ion. Iron ions play an important role in the body's cellular metabolism and are the core part of hemoglobin transporting oxygen. Most of the functional iron in the human body is present in the hemoglobin of circulating red blood cells. Some iron is present in myoglobin and many enzymes. The body's iron is stored in ferritin in cells, and iron circulates in the plasma bound to transferrin. Most of the iron that enters the circulation system is released by spleen and liver macrophages that are recycled from aging red blood cells. Iron deficiency and iron deficiency anemia are common nutritional and blood diseases. In infants and young children, the most common cause of iron deficiency is insufficient iron in the diet. In a rare case, iron deficiency anemia may be caused by mutations in TMPRSS6, a gene that encodes a membrane protease that normally acts as a transcriptional repressor of hepcidin, the main negative regulator of iron absorption. In young women, iron deficiency is usually caused by menstrual blood loss or blood loss during pregnancy, childbirth, and lactation. In the elderly, bleeding is often the main cause of iron deficiency and may originate from gastrointestinal bleeding, such as hemorrhoids, peptic ulcers, colon cancer, etc. Iron deficiency in infants can lead to impaired growth and intellectual development. Low serum ferritin concentration is a good indicator of iron deficiency, but ferritin levels increase due to inflammation and may be particularly high in cancer, macrophage activation syndrome, hepatitis or chronic kidney disease, which may mask the coexistence of iron deficiency and chronic inflammatory anemia. Plasma iron decreases and iron-binding capacity increases in severe iron deficiency, but these changes are not obvious in mild iron deficiency, and low plasma iron levels are also a feature of inflammatory anemia.

[0003] Currently, there is no conventional method and kit for detecting red blood cell iron in clinical practice. According to a 1962 document (Harold et al., New Method for the Determination of Whole Blood Iron and Hemoglobin, Clinical Chemistry), sodium hypochlorite can be used to release heme iron, and an iron color developer can be used to quantitatively analyze iron; however, during actual measurement, the applicant of the present invention found that sodium hypochlorite releases strong oxidizing gas (including chlorine), thereby causing environmental pollution and damaging the health of the experimenters; and the decomposition and release of sodium hypochlorite reduces the release efficiency of heme iron. Summary of the invention

[0004] The purpose of the present invention is to provide a method for releasing iron from red blood cells or bone marrow cells, a method for detecting iron content, and a kit. The present invention has developed a method for releasing iron from red blood cells or bone marrow cells (including red blood cells at different stages of red blood cell development), a method for detecting iron content, and a kit, which are helpful for releasing iron from red blood cells and bone marrow cells, as well as for detecting and diagnosing anemia.

[0005] In order to achieve the above objectives, the first aspect of the present invention discloses a method for releasing iron from red blood cells or bone marrow cells, wherein red blood cells or bone marrow cells are treated with an aqueous solution containing a surfactant and a nuclease to obtain a first treatment solution, the nuclease is used to digest and remove DNA in blood cells or bone marrow cells, and then an alkaline solution is added to the first treatment solution for incubation to obtain a second treatment solution, and then a sodium hypochlorite solution is added to the second treatment solution to release iron from red blood cells or bone marrow cells.

[0006] The reason for removing DNA from blood cells is that peripheral blood or other blood samples are usually used for erythrocyte iron release measurement, and the DNA of cells other than erythrocytes in the blood needs to be removed. Erythrocytes themselves do not contain DNA. Other components other than erythrocytes in peripheral blood do not affect iron detection, so there is no need to separate erythrocytes, and the peripheral blood can be directly tested.

[0007] In a preferred embodiment, the molar ratio of alkali to sodium hypochlorite is 1.1-3: 1. When the molar ratio of alkali to sodium hypochlorite is less than 1.1: 1, oxidizing gas will still be generated.

[0008] The applicant found that the reason for the large amount of chlorine released when sodium hypochlorite is used to release heme iron is the released iron ions, which accelerate the catalysis of sodium hypochlorite to produce chlorine. The decomposition of sodium hypochlorite can be inhibited by adding an excess of alkaline solution to prevent the release of harmful gases into the air, wherein sodium hydroxide and iron generate a small amount of water-insoluble iron hydroxide precipitation, which weakens the effect of iron ions catalyzing the decomposition of sodium hypochlorite (iron ions catalyze the decomposition of sodium hypochlorite).

[0009] The DNA in the cell nucleus is a viscous substance that is not completely soluble in water and may absorb some iron. The present invention digests the macromolecular DNA into fragments after adding nuclease digestion, which are completely soluble in water, thereby preventing DNA from absorbing iron and causing inaccurate iron content determination.

[0010] Furthermore, the aqueous solution containing the surfactant and the nuclease contains 20 U-800 U of nuclease (more preferably, 50 U-500 U of nuclease).

[0011] In a preferred embodiment, the volume ratio of the second treatment liquid to the sodium hypochlorite solution is 1-2:1-2 (more preferably, the volume ratio is 1:1), and the effective chlorine content in the sodium hypochlorite solution is 3-6%, and the percentage is the mass fraction.

[0012] Further, the volume ratio of the first treatment liquid to the alkaline solution is 1-2:1-2 (more preferably, the volume ratio is 1:1), and the concentration of the alkaline solution is 0.5-5 M (more preferably, the concentration of the alkaline solution is 2-5 M).

[0013] The second aspect of the present invention is to provide a method for determining the iron content in red blood cells or bone marrow cells, using the aforementioned method to release iron in red blood cells or bone marrow cells, then using an iron reducing agent to reduce trivalent iron ions to divalent iron ions, adding an iron ion color developer to develop color, and using an enzyme marker to detect the absorbance.

[0014] In a preferred embodiment, the iron reducing agent is ascorbic acid. Ascorbic acid reduces the ferric iron ions released from heme by the sodium hypochlorite solution to ferrous iron ions. A small amount of ferric hydroxide precipitate can react with the acidic ascorbic acid, and the iron re-enters the solution.

[0015] Sodium hydroxide is in excess, inhibiting the decomposition of sodium hypochlorite to prevent the release of harmful gases into the air. Ascorbic acid is subsequently added in an even greater excess to neutralize the remaining sodium hydroxide and sodium hypochlorite. After the sodium hypochlorite is neutralized, the generation of chlorine gas is further prevented.

[0016] Compared with acetic acid or sulfuric acid, ascorbic acid does not produce harmful gases such as chlorine when reacting with sodium hypochlorite.

[0017] The third aspect of the present invention is to disclose a red blood cell or bone marrow cell iron content detection kit, comprising an iron standard solution, an iron reducing agent, an iron color developing solution, a cell washing solution, an iron ion release promoting solution, a harmful gas inhibiting solution, and an iron releasing solution.

[0018] In a preferred embodiment, the iron reducing agent is ascorbic acid, the iron ion release promoting liquid is an aqueous solution containing a surfactant and a nuclease, the harmful gas suppressing liquid is an alkaline solution, and the iron releasing liquid is a sodium hypochlorite solution.

[0019] The fourth aspect of the present invention is to provide the use of the aforementioned red blood cell or bone marrow cell iron release method and the aforementioned red blood cell or bone marrow cell iron content determination method in the preparation of a kit for detecting the red blood cell or bone marrow cell iron content.

[0020] Compared with the prior art, the beneficial effects of the present invention are: The invention is safe, fast, accurate, easy to implement and low in cost, and is easy to popularize and use.

[0021] By adding excess alkaline solution, the decomposition of sodium hypochlorite can be inhibited to prevent the release of harmful gases into the air. Sodium hydroxide and iron generate a small amount of water-insoluble iron hydroxide precipitate, which weakens the effect of iron ions catalyzing the decomposition of sodium hypochlorite (iron ions catalyze the decomposition of sodium hypochlorite).

[0022] Ascorbic acid reduces the ferric iron ions released from hemoglobin by the sodium hypochlorite solution to ferrous iron ions without generating additional harmful gases. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 The test results of the iron standard solution (the color developing agent is o-phenanthroline), A. Gradient color development after adding iron color developing solution to iron standard solutions of different concentrations; B. OD values ​​of iron standard solutions of different concentrations detected by microplate reader; C. Draw a standard curve based on the concentration and OD value of the standard solution; Figure 2 . Detection of peripheral blood red blood cell iron content in mice with inflammatory anemia induced by bacterial lipopolysaccharide (endotoxin LPS) (the color developing agent is o-phenanthroline). (A) Peripheral blood red blood cells of LPS-injected mice and control group (injected with normal saline) mice, using the patented method for color development, where the first row is the negative control (i.e., o-phenanthroline is directly mixed with normal saline), and the second to fourth rows are 3 parallel samples of peripheral blood red blood cells of LPS-injected mice and control group (injected with normal saline); (B) OD values ​​of peripheral blood red blood cell iron detection of LPS-injected mice and control group (injected with normal saline) mice detected by microplate reader, where the first row is the negative control (i.e., o-phenanthroline is directly mixed with normal saline), and the second to fourth rows are 3 parallel samples of peripheral blood red blood cells of LPS-injected mice and control group (injected with normal saline); (C) Mouse peripheral blood iron content calculated according to the standard curve of the standard product (milligrams of iron contained in 1 ml of peripheral blood); (D) Peripheral blood red blood cell count of LPS-injected mice and control group (injected with normal saline) mice (detected by blood cell counter); Figure 3 Use starch potassium iodide test paper to develop color and detect the release of oxidizing gases; Figure 4 Use the chlorine gas detection tube to detect the release of chlorine gas. DETAILED DESCRIPTION

[0024] The present invention is described in detail below with reference to the accompanying drawings and specific embodiments.

[0025] Usually, sodium hypochlorite will not release a large amount of chlorine in a short time at room temperature and in the dark. However, in the detection process of using sodium hypochlorite to release iron ions, a large amount of chlorine is unexpectedly released in a short time at room temperature and in the dark. After research, the present invention finds that the reason for the large-scale release of oxidizing gas when using sodium hypochlorite to release heme iron is the released iron ions, which accelerate the catalysis of sodium hypochlorite to decompose and produce chlorine.

[0026] Take 0.1 ml of 0.5 mM ammonium ferric sulfate solution or ferric chloride solution, add 0.1 ml of water, add 0.2 ml of sodium hypochlorite (effective chlorine content 4%) solution, in a test tube, react for 3 minutes, and use starch potassium iodide test paper to detect. The test paper will turn blue, indicating that the reaction produces oxidizing gas. Therefore, it can be proved that the reason for the large amount of oxidizing gas released when sodium hypochlorite is used to release heme iron is the released iron ions, which accelerate the catalysis of sodium hypochlorite decomposition to produce chlorine.

[0027] The invention discloses a method for releasing iron from red blood cells or bone marrow cells, a method for detecting iron content, and a kit. First, an aqueous solution containing a surfactant (Triton X-100, deoxycholic acid, Tween-80, saponin, etc.) and a nuclease is used to treat red blood cells or bone marrow cells, and DNA is removed by nuclease digestion; then, an alkaline solution (sodium hydroxide, sodium carbonate, triethanolamine, pyridine solution, etc.) is added, and sodium hypochlorite is used to release the iron ions of heme in hemoglobin; then, ascorbic acid is used to reduce trivalent iron ions to divalent iron ions; an iron ion color developer (o-phenanthroline, ferrozine, phenanthroline, etc.) is added for color development, and an enzyme reader is used to detect absorbance; and the iron ion concentration in the sample is calculated according to a standard curve. Alkaline solutions such as sodium hydroxide can inhibit the decomposition of sodium hypochlorite to produce strong oxidizing harmful gases such as chlorine.

[0028] The red blood cell iron content detection kit comprises an iron standard solution, an iron reducing agent, an iron color developing solution, a red blood cell washing solution, an iron ion release promoting solution, a harmful gas suppressing solution and an iron releasing solution.

[0029] A bone marrow cell iron content detection kit comprises an iron standard solution, an iron reducing agent, an iron color developing solution, a cell washing solution, an iron ion release promoting solution, a harmful gas inhibiting solution, and an iron releasing solution.

[0030] Erythrocyte Iron Release: (1) Take 0.1 ml of anticoagulated blood (peripheral blood, or samples containing blood, from sources including humans, mice, rats, rabbits, etc.), centrifuge (1200-1500 rpm / 5-10 min), discard the supernatant, wash twice with saline, centrifuge, and discard the supernatant.

[0031] (2) Add about 0.5 ml of purified water containing a surfactant (Triton X-100, deoxycholic acid, Tween-80, saponin, etc.) and nuclease (50 U), mix well (using a pipette, ultrasonic mixing, or vortexing), and incubate at 15-37°C for 20 minutes (this is solution A).

[0032] (3) Take 0.1 ml of the above solution A, add 0.1 ml of sodium hydroxide solution (0.5-5 M sodium hydroxide solution, or sodium carbonate solution, triethanolamine, pyridine solution, etc.), mix well (blowing with a pipette or vortexing), and incubate at room temperature for 5 minutes (this is solution B).

[0033] (4) Take 0.1-0.2 ml of sodium hypochlorite solution (effective chlorine content 3-6%) and add it to Solution B. Mix well (using a pipette or vortexing), and incubate at room temperature for 15 minutes (this is Solution C).

[0034] Red blood cell iron content test: (1) Ascorbic acid: prepare a working solution with a concentration of 0.1-0.4 g / ml. Take 0.2 ml of ascorbic acid working solution and 0.2 ml of Solution C, mix well, and incubate at room temperature for 15 minutes. During this period, ultrasonic dissolution can be appropriately performed (this is Solution D).

[0035] (2) Take 50 μl of iron colorimetric solution (o-phenanthroline, ferrozine, phenanthroline, etc.) and add it to a 96-well plate. Add 150 μl of solution D and incubate at room temperature for 15-180 minutes. After the iron colorimetric solution and solution D are mixed, the absorbance can be measured at different time points, such as 15 minutes, 1 hour, 2 hours, and 3 hours.

[0036] (3) Measure the absorbance at 510 nm (for o-phenanthroline) or 562 nm (for phenanthroline). Calculate the sample concentration based on the standard curve.

[0037] The detection of iron release and iron content of bone marrow cells was performed as above, but with an increased amount of nuclease.

[0038] Ingredients of the red blood cell iron content detection kit: Iron standard solution: ammonium ferric sulfate or ferric chloride; Iron reducing agent: ascorbic acid; Iron colorimetric solution: o-phenanthroline or ferrozine or ferrozine; Red blood cell washing solution: 0.9% saline (or PBS and other buffer solutions); Iron ion release promoting solution: an aqueous solution containing a surfactant (containing 50 U of nuclease); Harmful gas inhibition solution: 3M sodium hydroxide solution (or sodium carbonate or triethanolamine or pyridine solution); Iron releasing solution: sodium hypochlorite solution.

[0039] Ingredients of bone marrow cell iron content detection kit: Iron standard solution: ammonium ferric sulfate or ferric chloride; Iron reducing agent: ascorbic acid; Iron colorimetric solution: o-phenanthroline or ferrozine or ferrozine; Cell washing solution: 0.9% saline (or PBS and other buffer solutions); Iron ion release promoting solution: aqueous solution containing surfactant (containing 500 U of nuclease); Harmful gas inhibition solution: 3M sodium hydroxide solution (or sodium carbonate or triethanolamine or pyridine solution); Iron releasing solution: sodium hypochlorite solution.

[0040] Example 1

[0041] Standard sample experimental steps (drawing standard curve): (1) Take the standard iron solution (0.1 M, ammonium ferric sulfate) and prepare 1 mM, 0.5 mM, 0.25 mM, 0.125 mM, 0.0625 mM, 0.03125 mM, and 0 mM iron standard solutions respectively.

[0042] (2) Take 100 μl of each of the above standards, add 100 μl of ascorbic acid (prepare ascorbic acid immediately before use, prepare 0.4 g / ml, and require ultrasonic dissolution), mix well, and react for 15 minutes.

[0043] (3) Take 50 μl of 20 mg / ml iron colorimetric solution (o-phenanthroline) and add it to a 96-well plate. Add 150 μl of the above reaction solution and incubate at room temperature for 15 minutes.

[0044] (4) Measure the absorbance at 510 nm and draw a standard curve.

[0045] The test results of the iron standard solution (the color developing agent is o-phenanthroline) are as follows Figure 1 shown. Figure 1 In A and B, from right to left, they correspond to 1 mM, 0.5 mM, 0.25 mM, 0.125 mM, 0.0625 mM, 0.03125 mM, and 0 mM, respectively. The concentrations in the same column are equal (i.e., 3 parallel samples).

[0046] Example 2

[0047] Erythrocyte Iron Release: (1) Take 0.1 ml of anticoagulated blood (mouse peripheral blood), centrifuge (1500 rpm / 10 min), discard the supernatant, wash twice with saline, centrifuge, and discard the supernatant. Peripheral blood is the peripheral blood of mice with inflammatory anemia induced by bacterial lipopolysaccharide (endotoxin LPS) (i.e., the mice were injected with LPS). Peripheral blood collection method: 0.5 mg / kg of mouse LPS was injected intraperitoneally twice a day for 8 consecutive days, and then the eyeballs were removed to collect blood and collect peripheral blood.

[0048] (2) Add about 0.5 ml of purified water containing a surfactant (Triton X-100, 1% (v / v)) and nuclease (50 U), mix well (using a pipette), and incubate at 37°C for 20 min (this is solution A).

[0049] (3) Take 0.1 ml of the above solution A, add 0.1 ml of sodium hydroxide solution (3 M sodium hydroxide solution), mix well (blowing evenly with a pipette), and incubate at room temperature for 5 minutes (this is solution B).

[0050] (4) Take 0.2 ml of sodium hypochlorite solution (effective chlorine content 4%) and add it to Solution B. Mix well (blowing evenly with a pipette), and incubate at room temperature for 15 minutes (this is Solution C).

[0051] Red blood cell iron content test: Ascorbic acid, prepare a working solution with a concentration of 0.4 g / ml, take 0.2 ml of ascorbic acid working solution, take 0.2 ml of solution C, mix well, incubate at room temperature for 15 minutes, and sonicate to assist dissolution (this is solution D).

[0052] (2) Take 50 μl of 20 mg / ml o-phenanthroline iron colorimetric solution and add it to a 96-well plate. Add 150 μl of solution D and incubate at room temperature for 15 minutes.

[0053] (3) Measure absorbance at 510 nm and calculate sample concentration based on the standard curve.

[0054] Comparative Example 1 It is basically the same as Example 2, except that the injected LPS is replaced by physiological saline.

[0055] Figure 2 . The iron content of peripheral blood red blood cells in mice with inflammatory anemia induced by bacterial lipopolysaccharide (endotoxin LPS) was detected (the color developing agent was o-phenanthroline). By comparing with the control group (injected with normal saline), the iron content of peripheral blood red blood cells of mice was reduced after LPS induction.

[0056] Example 3

[0057] Bone marrow cell iron test: (1) Take 0.1 ml of bone marrow cells (from mice), centrifuge (1500 rpm / 10 min), discard the supernatant, wash twice with saline, centrifuge, and discard the supernatant.

[0058] (2) Add about 0.5 ml of purified water containing a surfactant (Tween-80, 1% (v / v)) and nuclease (500 U), mix well (ultrasonic mixing), and incubate at 37°C for 20 min (this is solution A).

[0059] (3) Take 0.1 ml of the above solution A, add 0.1 ml of sodium hydroxide solution (3 M sodium hydroxide solution), mix well (blowing evenly with a pipette), and incubate at room temperature for 5 minutes (this is solution B).

[0060] (4) Take 0.2 ml of sodium hypochlorite solution (effective chlorine content 4%) and add it to Solution B. Mix well (blowing evenly with a pipette), and incubate at room temperature for 15 minutes (this is Solution C).

[0061] (5) Ascorbic acid: prepare a working solution with a concentration of 0.4 g / ml. Take 0.2 ml of ascorbic acid working solution and 0.2 ml of Solution C, mix well, incubate at room temperature for 15 min, and sonicate to assist dissolution (this is Solution D).

[0062] (6) Take 50 μl of iron colorimetric solution (20 mg / ml of o-phenanthroline) and add it to a 96-well plate. Add 150 μl of solution D and incubate at room temperature for 15 min.

[0063] (7) Measure absorbance at 510 nm. Calculate sample concentration based on the standard curve.

[0064] Comparative Example 2: Water Control Group The method is substantially the same as Example 3, except that the added sodium hydroxide solution is replaced by water.

[0065] like Figure 3 As shown, the starch potassium iodide test paper can make the test paper purple when detecting the water control group, indicating that the water control group produces oxidizing gas. However, the sodium hydroxide group (Example 3) does not make the test paper color, indicating that no oxidizing gas is produced. The starch potassium iodide test paper detection method is: take the above reaction C solution, add it to a test tube, put the moistened starch potassium iodide test paper at the mouth of the test tube, and react for about 1 minute.

[0066] like Figure 4As shown, the chlorine detection tube detected that the water control group had chlorine release, while the sodium hydroxide group (Example 3) had no chlorine release. The test results of starch potassium iodide test paper and chlorine detection tube indicate that excessive sodium hydroxide can prevent iron from catalyzing the release of chlorine from sodium hypochlorite, thereby proving that the experimental method invented by us is safe and feasible. The chlorine tube detection method is as follows: take the above reaction C liquid, add it to a test tube, connect the chlorine collector to the detection tube, extend the detection tube into the test tube mouth, and extract the gas.

[0067] The above embodiments are only used to illustrate the technical solutions of the present invention. Those skilled in the art should understand that the above embodiments do not limit the present invention in any form, and any technical solutions obtained by equivalent replacement or equivalent transformation shall fall within the protection scope of the present invention.

Claims

1. A method for releasing iron from red blood cells or bone marrow cells, characterized in that: Red blood cells or bone marrow cells are treated with an aqueous solution containing a surfactant and a nuclease to obtain a first treatment solution, and the nuclease is used to digest and remove DNA in the blood cells or bone marrow cells. Subsequently, an alkaline solution is added to the first treatment solution for incubation to obtain a second treatment solution, and then a sodium hypochlorite solution is added to the second treatment solution to release iron in the red blood cells or bone marrow cells.

2. The method for releasing iron from red blood cells or bone marrow cells according to claim 1, characterized in that: The volume ratio of the second treatment liquid to the sodium hypochlorite solution is 1-2:1-2, the effective chlorine content in the sodium hypochlorite solution is 3-6%, and the percentage is the mass fraction.

3. The method for releasing iron from red blood cells or bone marrow cells according to claim 1, characterized in that: The volume ratio of the first treatment liquid to the alkaline solution is 1-2:1-2, and the concentration of the alkaline solution is 0.5-5M.

4. The method for releasing iron from red blood cells or bone marrow cells according to claim 1, characterized in that: In the aqueous solution containing a surfactant and a nuclease, the amount of the nuclease is 20 U-800 U; and the volume percentage of the surfactant is 0.5-1.5%.

5. The method for releasing iron from red blood cells or bone marrow cells according to claim 1, characterized in that: The molar ratio of alkali to sodium hypochlorite is 1.1-3:

1.

6. A method for determining the iron content of red blood cells or bone marrow cells, characterized in that: The method according to any one of claims 1 to 5 is used to release iron in red blood cells or bone marrow cells, and then an iron reducing agent is used to reduce trivalent iron ions to divalent iron ions, an iron ion color developer is added to develop color, and an enzyme marker is used to detect the absorbance.

7. A method for determining the iron content of red blood cells or bone marrow cells according to claim 6, characterized in that: The iron reducing agent is ascorbic acid.

8. A kit for detecting the iron content of red blood cells or bone marrow cells, characterized in that: It includes iron standard solution, iron reducing agent, iron color developing solution, cell washing solution, iron ion release promoting solution, harmful gas inhibiting solution and iron releasing solution.

9. A kit for detecting iron content in red blood cells or bone marrow cells according to claim 8, characterized in that: The iron reducing agent is ascorbic acid, the iron ion release promoting liquid is an aqueous solution containing a surfactant and a nuclease, the harmful gas suppressing liquid is an alkaline solution, and the iron releasing liquid is a sodium hypochlorite solution.

10. Use of the method for releasing iron from erythrocytes or bone marrow cells according to any one of claims 1 to 5, or the method for determining the iron content in erythrocytes or bone marrow cells according to claim 6 or 7 in the preparation of a kit for detecting the iron content in erythrocytes or bone marrow cells.

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