Quality control product and preparation method for leukocyte differentiation antigen CD34 flow cytometry detection

Abstract

The present invention discloses a quality control product and a preparation method for flow cytometry detection of leukocyte differentiation antigen CD34, and relates to the technical field of quality control products for medical detection. The preparation method of the quality control product for flow cytometry detection of leukocyte differentiation antigen CD34 provided by the present invention, by adopting microspheres coupled with CD34 and CD45 proteins to replace genetically engineered cells, and utilizing the physical stability and uniform particle size distribution of the microsphere material, significantly improves the consistency of quality control products between batches, while extending the shelf life of the product. Furthermore, in the peripheral blood processing link, by optimizing the sequence of first fixing the peripheral blood cells, then adding back the plasma, and finally mixing with the microspheres, it helps to stabilize the cell membrane structure and effectively avoids cell rupture and the release of endogenous interfering substances. The optimization of this sequence makes the quality control product closer to the actual processing conditions of clinical samples, which not only improves the specificity and signal-to-noise ratio of the flow detection signal, but also enhances the correlation between the quality control results and actual clinical detection.

Description

Technical Field

[0001] The present invention relates to the technical field of quality control products for medical detection, and in particular to a quality control product for flow cytometry detection of leukocyte differentiation antigen CD34 and a preparation method thereof. Background Art

[0002] Hematopoietic stem cell transplantation can restore hematopoietic function with minimal tumor cell contamination, and is widely used to treat a variety of malignant blood diseases, solid tumors, genetic diseases, and severe combined immunodeficiency. Hematopoietic stem cells are primarily derived from peripheral blood, bone marrow, and umbilical cord blood, with umbilical cord blood being the most abundant source. Umbilical cord blood is increasingly recognized and valued by physicians and patients due to its plentiful source, ease of collection, and increasingly simplified storage and transplantation techniques.

[0003] Currently, the important indicators for evaluating hematopoietic stem cell transplantation include the total number of nucleated cells and CD34 + The total number of cells. Among them, CD34 + The number and functional status of cells directly determine transplant efficacy. This is primarily measured using flow cytometry using a single platform (absolute count) or a dual platform (percentage count combined with a complete blood count). Current testing systems rely on standardized quality control products for quality control to ensure data accuracy and interlaboratory comparability.

[0004] At present, mainstream quality control products are mainly imported, and the main manufacturers are BD, Beckman Coulter, R&D and Streck. However, the high cost, complicated transportation process and short shelf life of imported quality control products after arrival have limited their widespread application, making it difficult for some laboratories to achieve normalized quality control management. The comparability of cross-laboratory results is poor and the quality is uncontrollable, which affects the judgment and efficacy of clinical hematopoietic stem cell transplantation. Although there are technologies in China that attempt to develop quality control products through genetic engineering to construct cell lines, there are CD34 + Deficiencies such as high discreteness of cell population distribution and side scatter signal (SSC) offset further highlight the technical bottlenecks of the existing quality control system.

[0005] Domestic and international technical regulations impose strict requirements on stem cell testing processes and quality control, particularly setting clear standards for cell counting accuracy and stability. However, the performance limitations of existing quality control products not only increase the risk of test result fluctuations but may also affect clinical scientific assessments of transplant outcomes. Therefore, developing a quality control product that can optimize cell distribution characteristics and enhance test stability has become a key requirement for improving hematopoietic stem cell testing systems. Summary of the Invention

[0006] The technical problem to be solved by the present invention is the CD34 + To solve the problems of high discreteness of cell population distribution and large side scatter signal (SSC), a quality control product, preparation method and application for flow cytometry detection of leukocyte differentiation antigen CD34 are provided.

[0007] In order to solve the above problems, the present invention proposes the following technical solutions:

[0008] In a first aspect, the present invention provides a method for preparing a quality control product for flow cytometry detection of leukocyte differentiation antigen CD34, comprising the following steps:

[0009] S1. Take 20 μL-200 μL of microspheres, wash and activate them, then add 10-500 μL of CD34 protein and 10-500 μL of CD45 protein for incubation. After incubation, add blocking agent. Wash the blocked microspheres and store them at 4°C to 8°C to obtain CD34 microspheres.

[0010] S2. Collect peripheral blood from healthy adults and separate plasma and blood cells;

[0011] S3, fixing the blood cells and adding the plasma to obtain fixed peripheral blood;

[0012] S4, adding the CD34 microspheres obtained in S1 to the peripheral blood obtained in S3 according to a preset ratio, and mixing thoroughly to obtain a quality control product;

[0013] S5. Determine the absolute number of CD34 microspheres and white blood cells in the quality control product obtained in S4.

[0014] A further technical solution is that the step S3 further includes counting the white blood cells in the fixed peripheral blood, so that the white blood cell concentration in the fixed peripheral blood is 4.0×10 9 ~ 10.0×10 9 cells / L.

[0015] A further technical solution is that the operation of fixing the blood cells includes:

[0016] Prepare 0.005-0.75% (w / v) CrCl3 solution and 0.1-0.5% (w / v) paraformaldehyde solution in advance using solvent;

[0017] The CrCl3 solution and the blood cells are mixed in a volume ratio of 1:1, fixed at 0~8℃ for 5min~18h, centrifuged to remove the supernatant, and washed with PBS to remove the CrCl3 solution; then, the paraformaldehyde solution and the blood cells are mixed in a volume ratio of 1:1, fixed at 0℃~8℃ for 16-22h, centrifuged to remove the supernatant, and washed with PBS to remove the paraformaldehyde solution to obtain fixed blood cells.

[0018] Its further technical solution is that in step S4, the preset ratio includes: + In low-value quality control products, the proportion of CD34 microspheres in peripheral blood leukocytes is 0.1% to 0.3%; + In high-value quality control products, the proportion of CD34 microspheres in peripheral blood leukocytes is 0.3% to 0.6%.

[0019] A further technical solution is that the microsphere washing operation includes: taking 20 μL-200 μL of microspheres, adding 1-5 mL of MES coupling buffer, mixing, rotating and incubating for 5-30 minutes, centrifuging to remove the supernatant, repeating the washing 1-3 times, and resuspending the microspheres in MES coupling buffer.

[0020] A further technical solution is that the microsphere activation operation includes: adding a reaction solution containing 10 μL-200 μL EDC and 10 μL-200 μL NHS to the cleaned microspheres, mixing, and incubating with rotation at 37°C for 10-30 min.

[0021] A further technical solution is that in step S5, a flow cytometer is used to measure the CD34 microsphere ratio of the quality control product and determine the target value, and a five-category blood cell counter is used to measure the absolute number of white blood cells in the quality control product and determine the target value.

[0022] A further technical solution is that the microspheres are selected from silica carboxyl microspheres, polystyrene microspheres or magnetic microspheres, and the particle size of the microspheres is 3-20 μm, preferably 5-15 μm, and more preferably 5-13 μm.

[0023] In a second aspect, the present invention provides a quality control product for flow cytometry detection of leukocyte differentiation antigen CD34, which is prepared by the method described in the first aspect.

[0024] The present invention also provides the quality control product for flow cytometry detection of leukocyte differentiation antigen CD34, or the quality control product prepared by the preparation method of the quality control product for flow cytometry detection of leukocyte differentiation antigen CD34 in the first aspect. + Application in cell kits.

[0025] Compared with the prior art, the present invention can achieve the following technical effects:

[0026] The method for preparing a quality control product for flow cytometry testing of the leukocyte differentiation antigen CD34, provided herein, avoids the limitations of traditional techniques that rely on complex cell culture and genetic modification by using microspheres coupled to CD34 and CD45 proteins instead of genetically engineered cells. This directed coupling of proteins to the microsphere surface not only simplifies the preparation process and reduces biosafety risks, but also significantly improves batch-to-batch consistency of the quality control product through the physical stability and uniform particle size distribution of the microsphere material, while also extending the product's shelf life and addressing the performance instability associated with traditional genetically engineered cells due to activity decay or epitope variation.

[0027] In addition, the preparation method of the present invention optimizes the order of first fixing the peripheral blood cells, then adding back the plasma, and finally mixing with the microspheres in the peripheral blood processing link, thus breaking through the defect of direct mixing and then fixing in the prior art that easily damages the cells. Completing the fixation of blood cells before mixing can reduce the impact of the solutions and operations used in the mixing process on the proteins on the surface of the microspheres, avoiding pH changes causing protein denaturation or frequent operations reducing the amount of protein on the surface of the microspheres. This optimization of the sequence maintains the stability of the quality control product, which not only improves the specificity and signal-to-noise ratio of the flow detection signal, but also enhances the correlation between the quality control results and actual clinical detection.

[0028] In summary, this method overcomes the limitations of traditional quality control products CD34 while retaining the biomimetic properties of clinical samples through the collaborative innovation of microsphere coupling and processing sequence. + Technical bottlenecks such as cell dispersion, high SSC and short shelf life provide CD34 flow cytometry with standardized quality control products that are more stable, easier to operate and have wider clinical applicability. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the following briefly introduces the drawings required for use in the description of the embodiments. Obviously, the drawings described below are some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0030] Figure 1 A flow chart of a method for preparing a quality control product for flow cytometry detection of leukocyte differentiation antigen CD34 provided in an embodiment of the present invention;

[0031] Figure 2 The stability test results of the low-value quality control and high-value quality control prepared in the embodiment of the present invention;

[0032] Figure 3 This is the test result of the control group of low-value quality control products produced by BD;

[0033] Figure 4The test results of the low-value quality control product experimental group produced by BD;

[0034] Figure 5 This is the test result of the control group of high-value quality control products produced by BD;

[0035] Figure 6 Test results of the high-value quality control product experimental group produced by BD;

[0036] Figure 7 This is the test result of the KRC2201 low-value quality control group;

[0037] Figure 8 This is the test result of the KRC2201 low-value quality control product experimental group;

[0038] Figure 9 This is the test result of the control group of KRC2201 high-value quality control product;

[0039] Figure 10 This is the test result of the KRC2201 high-value quality control product experimental group;

[0040] Figure 11 The test results of the low-value quality control group prepared in this example are as follows;

[0041] Figure 12 The test results of the low-value quality control product experimental group prepared in this embodiment;

[0042] Figure 13 The test results of the high-value quality control group prepared in this example;

[0043] Figure 14 These are the test results of the high-value quality control product experimental group prepared in this example. DETAILED DESCRIPTION

[0044] The present invention is further illustrated below by way of specific application examples, but the invention is not limited to these examples. Experimental methods not specifying specific conditions in the following application examples should be performed according to conventional methods and conditions, or according to the product specifications. Unless otherwise specified, the reaction temperature generally refers to room temperature, which in this invention refers to 16°C to 30°C.

[0045] Unless otherwise specified, the experimental methods used in the following application examples are conventional methods.

[0046] Unless otherwise specified, the materials and reagents used in the following application examples can be purchased from commercial sources. The CD34 protein and CD45 protein used in the examples are commercially available recombinant human CD34 protein and human recombinant CD45 protein, both of which are His-tagged proteins expressed in HEK293 cells.

[0047] The present invention provides a method for preparing a quality control product for flow cytometry detection of leukocyte differentiation antigen CD34, comprising the following steps:

[0048] S1. Take 20 μL-200 μL of microspheres, wash and activate them, then add 10-300 μL of CD34 protein and 10-300 μL of CD45 protein for incubation. After incubation, add blocking agent. Wash the blocked microspheres and store them at 4°C to 8°C to obtain CD34 microspheres.

[0049] S2. Collect peripheral blood from healthy adults and separate plasma and blood cells;

[0050] S3, fixing the blood cells and adding the plasma to obtain fixed peripheral blood;

[0051] S4, adding the CD34 microspheres obtained in S1 to the peripheral blood obtained in S3 according to a preset ratio, and mixing thoroughly to obtain a quality control product;

[0052] S5. Determine the absolute number of CD34 microspheres and white blood cells of the quality control product obtained in S4.

[0053] See further Figure 1 , which is a flow chart of a method for preparing a quality control product for flow cytometry detection of leukocyte differentiation antigen CD34 provided in an embodiment of the present invention.

[0054] The following details each step:

[0055] S11, cleaning of silica carboxyl microspheres

[0056] Take 40 μL of 5 μm diameter silica carboxyl microspheres and add 1 mL of coupling buffer MES (50 mM, pH 6.0). Mix well by pipetting. Incubate with rotation at room temperature for 10 min. Remove the supernatant by centrifugation (6000 rpm, 10 min). Repeat the washing once. Finally, resuspend the silica carboxyl microspheres in 500 μL of coupling buffer.

[0057] S12, Activation of Silica Carboxyl Microspheres

[0058] Slowly add 20 μL of freshly prepared 50 mg / mL EDC and 20 μL of 50 mg / mL NHS reaction solution to the centrifuge tube, mix well by pipetting, and incubate with rotation at 37°C for 15 min.

[0059] S13, Microspheres and Protein Coupling

[0060] After incubation, the supernatant was removed by centrifugation, and the tube was washed once with 1 mL of MES buffer. 80 µL of 1 mg / mL CD34 protein and 40 µL of 1 mg / mL CD45 protein were added, and the tube was incubated with rotation at 37 °C for 2 hours.

[0061] S14, microsphere surface group sealing

[0062] After the coupling incubation is completed, the supernatant is removed by centrifugation, and then 500 μL of 2% BSA blocking solution is added to the centrifuge tube for blocking, and the tube is rotated and incubated at 37°C for 1 hour.

[0063] S15. Microsphere cleaning and storage

[0064] After blocking, the microspheres were centrifuged and the supernatant was removed. The microspheres were washed once with 1 mL of blocking solution (2% BSA), and then resuspended in 500 μL of PBS buffer and refrigerated at 4°C to 8°C.

[0065] S21. Human peripheral blood collection

[0066] Use EDTA as the anticoagulant blood collection tube to collect 90 mL of adult peripheral blood, transfer the peripheral blood into a mold-free, sterile, and pyrogen-free 250 mL preparation bottle and mix thoroughly.

[0067] S22. Centrifugal separation of plasma and blood cells

[0068] After mixing the peripheral blood, take 40 mL and transfer them to two 50 mL centrifuge tubes, mark the low value and high value respectively, centrifuge at 4°C and 2000g for 10 min, aspirate the plasma corresponding to the low value and high value, and refrigerate at 2-8°C for future use.

[0069] S31. Blood cell fixation

[0070] 24 h before fixation, 0.1% (w / v) CrCl3 solution and 0.35% (w / v) paraformaldehyde solution were prepared using DPBS.

[0071] Mix the CrCl₃ solution with the centrifuged peripheral blood cells at a 1:1 volume ratio and fix at 2–8°C for 1 h. Centrifuge at 2000 g for 10 min at 4°C, remove the supernatant, and wash the cells twice with DPBS, which is then discarded. Then, mix the paraformaldehyde solution with the centrifuged peripheral blood cells at a 1:1 volume ratio and fix at 2–8°C for 18 h. Centrifuge at 2000 g for 10 min at 4°C, remove the supernatant, and wash the cells twice with DPBS, which is then discarded. This fixated blood cells are then obtained.

[0072] The low-value and high-value plasma were added back to the corresponding fixed blood cells and mixed to obtain fixed peripheral blood (low-value, high-value).

[0073] S32, human peripheral blood leukocyte count

[0074] 2 mL of fixed high and low peripheral blood were collected and the white blood cells were counted 15 times in a row using a five-differential blood cell counter. The average white blood cell density of the 15 results was calculated as follows: low value: 7.734×10 9 cells / L, CV: 1.59%; high value: 7.681×10 9 cells / L, CV: 1.70%.

[0075] 30 μL of protein-coupled microspheres were added to 1 mL of PBS solution without adding cell dye. The average microsphere concentration was measured by trypan blue counting method and was 3.900 × 10 6 pieces / mL.

[0076] S41, microspheres mixed with peripheral blood cells

[0077] According to the low mean white blood cell density of 7.734×10 9 cells / L and microsphere density 3.900×10 6 cells / mL, the residual volume of peripheral blood was 38 mL, the percentage of CD34 microspheres in the total white blood cells in the low-value quality control was 0.100%; the high-value average white blood cell density was 7.681×10 9 cells / L and microsphere density 3.900×10 6 / mL, the residual volume of peripheral blood is 38 mL, the percentage of CD34 microspheres in the total leukocytes in the high-value quality control is 0.450%, the amount of microspheres added to the low-value and high-value quality controls are calculated and added respectively, and after thorough mixing, they are stored at 2-8°C to obtain low-value quality control products and high-value quality control products.

[0078] S51, CD34 microspheres and absolute leukocyte counts

[0079] Take the above CD34 + 2.5 mL of low-value and high-value quality control samples were used to count leukocytes 20 times using a five-differential hematology counter. The average white blood cell density of these 20 results was calculated as follows: low-value quality control: 7760 cells / μL, CV: 1.59%; high-value quality control: 7681 cells / μL, CV: 1.70%.

[0080] The CD34 microsphere proportions of low-value and high-value quality controls were tested 20 times in a row using flow cytometry, and the average CD34 microsphere proportions of the 20 results were calculated as follows: low-value quality control: 0.111% (0.089-0.132%); high-value quality control: 0.490% (0.447-0.532%).

[0081] The above embodiment uses silica carboxyl microspheres as an example to introduce the quality control product for flow cytometry detection of leukocyte differentiation antigen CD34 and its preparation method. In other embodiments, those skilled in the art may also use polystyrene microspheres or magnetic microspheres, etc., and the present invention is not limited thereto.

[0082] Experiment 1: Stability Test

[0083] The 120-day continuous test method (0 days, 3 days, 10 days, 14 days, 18 days, 24 days, 31 days, 38 days, 45 days, 50 days, 65 days, 70 days, 75 days, 80 days, 85 days, 90 days, 95 days, 100 days, 105 days, 111 days, 115 days, 120 days) was used to test the changes in the proportion of low-value quality control and high-value quality control CD34 microspheres prepared in this embodiment. The test results are shown in Figure 2 It can be seen that the test values from day 0 to day 120 are all within the target value range (low value quality control: 0.089-0.132%, high value quality control: 0.447-0.532%). Therefore, the quality control product prepared in this embodiment of the present invention can be stably stored within 120 days, and the validity period of the quality control can reach 120 days.

[0084] Test Method: a. Prepare two flow cytometer tubes, one labeled as the isotype control tube and the other as the experimental tube, for each CD34 control blood sample. Add 100 μL of each CD34 control blood sample to each tube. b. Add 10 μL each of FITC-CD45 and PE-ISO antibodies to the control tube and 10 μL each of FITC-CD45 and PE-CD34 antibodies to the experimental tube. Gently shake to mix thoroughly, then incubate at 4°C in the dark for 30 min. c. After incubation, remove the control blood sample and add 1 mL of 1× hemolysin to each tube, mix thoroughly, and allow to hemolyze in a 4°C refrigerator in the dark for 10 min. d. Remove the control blood sample and centrifuge at 500 g for 3 min. Discard the supernatant, resuspend in 1 mL of 1× PBS, mix thoroughly, and centrifuge at 500 g for 3 min twice. e. Discard the supernatant, resuspend in 300-500 μL of 1× PBS, mix thoroughly, and prepare for analysis.

[0085] The quality control product prepared by the embodiment of the present invention was compared with the quality control product produced by BD (product name: Stem CellControl CD34 + The performance of the product (Whole Blood Process Control) and Shenzhen Kenuo Medical Laboratory's product (KRC2201) were compared.

[0086] The test results of the quality control products produced by BD are shown in Figure 3-Figure 6 ; KRC2201 quality control product test results are shown in Figure 7-10 ; The quality control product prepared in the embodiment of the present invention is shown in Figure 11-14 .

[0087] The flow cytometry graph shows that in the low-value experimental group of our company's previous generation product KRC2201, CD34 + The cell distribution was obviously dispersed and the SSC was significantly higher, indicating that the low-value quality control product was not stable enough and its performance did not meet expectations.

[0088] In the KRC2201 high value experimental group, CD34 + The aggregation of the cell population is slightly better than that of the low-value group, but the SSC is still too high, indicating that the performance of the high-value quality control product is still unstable.

[0089] The low value experimental group of the quality control product of the embodiment of the present invention showed that CD34 + The cell population distribution is highly concentrated and the SSC value is significantly reduced, indicating that the performance control is better than that of the KRC2201 product and the SSC value remains in the ideal range.

[0090] In the high-value experimental group, the CD34 + The cell population remained tightly aggregated and the SSC value was significantly reduced, indicating that its high-value quality control product has excellent stability.

[0091] The low-value quality control product and high-value quality control product provided by the embodiment of the present invention, CD34 + The cells are concentrated and the SSC value is stable. Compared with the imported BD company's quality control products and KRC2201 products, the CD34 in the embodiment of the present invention is + The cells are clearly clustered and easier to gate, which can significantly reduce the differences in gating between different experimenters. The comprehensive performance surpasses the KRC2201 product in all aspects, and is also better than the imported BD company's quality control products in terms of shelf life.

[0092] The above results show that the quality control product provided by the present invention significantly improves CD34 + The centralized cell distribution and effective reduction of the SSC value solve the key problems of the previous generation product (KRC2201) and provide better performance. In addition, the quality control product provided by the present invention has a shelf life of up to 120 days and significantly improved stability.

[0093] In the above embodiments, the description of each embodiment has its own focus. For parts that are not described in detail in a certain embodiment, reference can be made to the relevant descriptions of other embodiments.

[0094] The above description is a specific embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any person skilled in the art can easily conceive of various equivalent modifications or substitutions within the technical scope disclosed in the present invention, and such modifications or substitutions are intended to be within the scope of protection of the present invention. Therefore, the scope of protection of the present invention shall be subject to the scope of protection of the claims.

Claims

1. A method for preparing a quality control product for flow cytometry detection of leukocyte differentiation antigen CD34, characterized in that: The following steps are involved: S1. Take 20 μL-200 μL of microspheres, wash and activate them, then add 10-500 μL of CD34 protein and 10-500 μL of CD45 protein for incubation. After incubation, add blocking agent. Wash the blocked microspheres and store them at 4°C to 8°C to obtain CD34 microspheres. S2. Collect peripheral blood from healthy adults and separate plasma and blood cells; S3, fixing the blood cells and adding the plasma to obtain fixed peripheral blood; S4, adding the CD34 microspheres obtained in S1 to the peripheral blood obtained in S3 according to a preset ratio, and mixing thoroughly to obtain a quality control product; S5. Determine the absolute number of CD34 microspheres and white blood cells in the quality control product obtained in S4; Wherein, the operation of fixing the blood cells includes: Prepare 0.005-0.75% w / v CrCl3 solution and 0.1-0.5% w / v paraformaldehyde solution in advance using solvent; The CrCl3 solution and the blood cells are mixed in a volume ratio of 1:1, fixed at 0~8℃ for 5min~18h, centrifuged to remove the supernatant, and washed with PBS to remove the CrCl3 solution; then, the paraformaldehyde solution and the blood cells are mixed in a volume ratio of 1:1, fixed at 0℃~8℃ for 16-22h, centrifuged to remove the supernatant, and washed with PBS to remove the paraformaldehyde solution to obtain fixed blood cells.

2. The method for preparing a quality control product for flow cytometry detection of leukocyte differentiation antigen CD34 according to claim 1, wherein: The step S3 also includes counting the white blood cells in the fixed peripheral blood, so that the white blood cell concentration in the fixed peripheral blood is 4.0×10 9 ~ 10.0×10 9 cells / L.

3. The method for preparing a quality control product for flow cytometry detection of leukocyte differentiation antigen CD34 according to claim 1, wherein: In step S4, the preset ratio includes: + In low-value quality control products, the proportion of CD34 microspheres in peripheral blood leukocytes is 0.1% to 0.3%; + In high-value quality control products, the proportion of CD34 microspheres in peripheral blood leukocytes is 0.3%~0.6%.

4. The method for preparing a quality control product for flow cytometry detection of leukocyte differentiation antigen CD34 according to claim 1, wherein: The microsphere washing operation includes: taking 20 μL-200 μL of microspheres, adding 1-5 mL of MES coupling buffer, mixing, rotating and incubating for 5-30 minutes, centrifuging to remove the supernatant, repeating the washing 1-3 times, and resuspending the microspheres in MES coupling buffer.

5. The method for preparing a quality control product for flow cytometry detection of leukocyte differentiation antigen CD34 according to claim 1, wherein: The microsphere activation operation includes: adding a reaction solution containing 10 μL-200 μL of EDC and 10 μL-200 μL of NHS to the washed microspheres, mixing, and incubating at 37° C. with rotation for 10-30 minutes.

6. The method for preparing a quality control product for flow cytometry detection of leukocyte differentiation antigen CD34 according to claim 1, wherein: In step S5, a flow cytometer is used to measure the CD34 microsphere ratio of the quality control product and determine the target value, and a five-category blood cell counter is used to measure the absolute number of white blood cells of the quality control product and determine the target value.

7. The method for preparing a quality control product for flow cytometry detection of leukocyte differentiation antigen CD34 according to claim 1, wherein: The microspheres are selected from silica carboxyl microspheres, polystyrene microspheres or magnetic microspheres, and the particle size of the microspheres is 3-20 μm.

8. A quality control product for flow cytometry detection of leukocyte differentiation antigen CD34, characterized in that: Prepared by the method according to any one of claims 1 to 7.

9. The quality control product for flow cytometry detection of leukocyte differentiation antigen CD34 according to claim 8 is used in the preparation of the CD34 detection + Application in cell kits.

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