Antibody combination for detecting minimal residual disease and application thereof

Abstract

The invention discloses an antibody combination for detecting minimal residual disease and application thereof. The antibody combination comprises a CD45 antibody, a CD19 antibody, a CD33 antibody, a CD10 antibody, a CD20 antibody, a CD38 antibody, a CD34 antibody, a CD58 antibody, a CD66c antibody, a CD73 antibody, a CD81 antibody, a CD123 antibody and a CD304 antibody. A specific antibody combination for detecting the minimal residual disease is designed, a specific gate setting strategy is combined, 13-color flow analysis is realized, and target cells can be accurately locked and normal cells and differential cells can be effectively distinguished by combining composition and development process analysis of CD19 + cells and leukemia cell related immunophenotype (LAIP) analysis; the detection rate, the accuracy and the sensitivity are obviously improved.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of biomedical technology, and relates to an antibody combination for detecting minimal residual disease and application thereof. BACKGROUND

[0002] Minimal residual disease (MRD) refers to a state that a small amount of tumor cells remains in the body of a blood system tumor patient after complete remission is obtained. At present, due to the application of multiple drugs in combination, most blood disease patients can obtain morphological complete remission. However, some patients still relapse, and the main reason is that a small amount of morphologically indistinguishable residual tumor cells exist in the body of the patient. Regular detection of MRD is part of the detection of treatment response, which can judge the sensitivity of abnormal cells to chemotherapeutic drugs and the degree of bone marrow regeneration during treatment. In particular, the patient's MRD is continuously positive, the MRD level gradually increases, and the MRD is converted from negative to positive, which has the significance of differentiating high-risk recurrence and poor prognosis, and effectively provides help for clinical prognosis judgment and treatment.

[0003] The identification of normal B precursor cells and leukemia cells has been a problem in judging the remission degree of B-ALL (B lymphoblastic leukemia). After chemotherapy and stem cell transplantation in normal children or adults, a considerable number of normal B precursor cells can appear in the bone marrow during the recovery period, and it has been reported that the number can reach 50% of nucleated cells. If only morphological criteria are used, it may be judged as relapse. Due to the gradual deepening of the understanding of this group of cells in recent years, especially the in-depth understanding of the characteristics of their immunophenotype, and the application of multi-color flow cytometry, the development process of normal B cells can be more clearly displayed, so that accurate detection of B-ALL MRD becomes possible.

[0004] The analysis method used for detecting residual leukemia cells by using multi-parameter flow cytometry mainly monitors leukemia-associated immunophenotype (LAIP) or "different from normal expression pattern" (DFN). The LAIP method needs to screen the indicators at the time of initial diagnosis immunotyping, otherwise it is easy to miss diagnosis. The DFN method has high requirements for the experience and professional knowledge level of the analysis personnel. Multi-parameter flow cytometry analysis can be detected by using a traditional flow cytometer, but it is affected by the detection channel. At present, a 6-10 color detection scheme is mostly used. Each sample needs to be detected by multiple different detection methods. Not only the skeleton antibody needs to be repeatedly detected, but also a large amount of sample is required, and the sensitivity is not high.

[0005] In summary, how to provide a detection technology with high sensitivity, high accuracy, high detection rate and simple and rapid detection is one of the problems to be solved in the field of MRD detection. SUMMARY

[0006] In view of the deficiencies of the prior art and actual needs, the application provides an antibody combination for detecting minimal residual disease and application thereof, designs a specific antibody combination, combines composition and development analysis of CD19+ cells and analysis ideas of leukemia cell-related immunophenotype (LAIP), and significantly improves detection rate, sensitivity and accuracy.

[0007] To achieve the above object, the application adopts the following technical solutions.

[0008] In a first aspect, the application provides an antibody combination for detecting minimal residual disease, which comprises CD45 antibody, CD19 antibody, CD33 antibody, CD10 antibody, CD20 antibody, CD38 antibody, CD34 antibody, CD58 antibody, CD66c antibody, CD73 antibody, CD81 antibody, CD123 antibody and CD304 antibody.

[0009] In the application, a specific antibody combination for detecting minimal residual disease is designed, the analysis of composition and development process (i.e. DFN different from normal) of CD19+ cells is combined with the analysis of leukemia cell-related immunophenotype (LAIP), the situation of immunophenotype change after leukemia treatment is considered, and the detection rate, sensitivity and accuracy can be significantly improved.

[0010] Preferably, the antibodies in the antibody combination all contain fluorescein labels.

[0011] Preferably, the CD45 antibody, CD19 antibody, CD33 antibody, CD10 antibody, CD20 antibody, CD38 antibody, CD34 antibody, CD58 antibody, CD66c antibody, CD73 antibody, CD81 antibody, CD123 antibody and CD304 antibody are sequentially labeled with V500, BV780, PECY7, APC-CY7, ECD, BV421, PercP-cy5.5, APC, FITC, PE, BV660, APC-A700 and BV610.

[0012] In the application, a specific fluorescein labeling strategy is designed for the specific antibody combination, which can reduce spectral overlap, so as to better adapt to subsequent thirteen-color flow cytometry analysis and detection process.

[0013] Preferably, the antibodies in the antibody combination are monoclonal antibodies.

[0014] In the application, the antibody itself is well known to those skilled in the art, which specifically binds (anti) corresponding antigen.

[0015] In a second aspect, the present application provides use of the antibody combination for detecting minimal residual disease in the manufacture of a product for detecting minimal residual disease.

[0016] In a third aspect, the present application provides a kit for detecting minimal residual disease, the kit comprising the antibody combination for detecting minimal residual disease of the first aspect.

[0017] Preferably, the kit further comprises any one or a combination of at least two of a cell lysis solution, a membrane disrupter, a buffer, or a flow tube for use with a flow cytometer.

[0018] In a fourth aspect, the present application provides an apparatus for detecting minimal residual disease, the apparatus comprising a detection unit and an analysis unit.

[0019] The detection unit is configured to perform operations comprising:

[0020] The sample to be tested is stained with the antibody combination for detecting minimal residual disease of the first aspect, and flow cytometry is performed.

[0021] The analysis unit is configured to perform operations comprising:

[0022] The cells are gated, and normal cells and abnormal cells are distinguished based on the expression profile of the antibody combination.

[0023] Preferably, the instrument for flow cytometry comprises a thirteen-color flow cytometer.

[0024] Preferably, the thirteen-color flow cytometer comprises a three-laser thirteen-color flow cytometer of Beckman Coulter DxFLEX.

[0025] Preferably, the flow cytometry is performed to obtain a cell number of 2,000,000-5,000,000, including but not limited to 2,500,000, 3,000,000, 3,500,000, 4,500,000, 4,600,000, 4,800,000, or 4,900,000.

[0026] The traditional detection method (such as 8-10 color flow) may have the problems of insufficient sensitivity or incomplete phenotype coverage, while the present application successfully designs a 13-color one-tube detection scheme for the first time, designs specific antibody combinations, and creatively combines the use of 3-laser 13-color flow cytometry design analysis scheme to make up for the market gap. Through specific marker combinations, the antibody combination design selects B cell development stage markers: CD19, CD20, CD10, CD34, CD38, CD45 (to distinguish normal B progenitor cells and leukemia cells); abnormal expression markers: CD33 (myeloid marker, may be abnormally expressed in B-ALL), CD123 (common in leukemia stem cells), CD66c (highly expressed in early B-ALL), CD73, CD304 (heterogeneous expression in B-ALL), CD81 (weak expression in B-ALL), and enhances the recognition ability of abnormal cells, especially suitable for the detection of low-level MRD (such as <0.01%), and realizes high sensitivity, accuracy, detection rate and simple and rapid detection.

[0027] The 13-color one-tube detection scheme designed in the present application can be used for disease diagnosis purposes and non-disease diagnosis purposes, such as distinguishing different sample sources, drug screening, and basic mechanism research, etc.

[0028] Preferably, the gating method comprises:

[0029] Sequentially removing cell adhesion by FSC-A and FSC-H double parameters; selecting live cell gate by FSC-SSC; setting blood cell gate by CD45-SSC sequential gating method; setting gate by CD19-SSC reverse method, which is used for circle gate analysis of all B-lineage cells (including normal cells and abnormal cells); finally, back gate setting, which is used for judging the true and false of cell population by reversing the setting sequence;

[0030] Using CD19 gating method and CD45 gating method, the CD19 gating method is used for circle gate analysis of all B-lineage cells (including normal cells and abnormal cells), and the cell source is determined according to the antibody expression; the CD45 gating method is used to distinguish various types of cells in the sample, including lymphocytes, granulocytes, monocytes, blast cells and nucleated red blood cells.

[0031] Specifically, the enhancement of CD19 expression and the weakening of CD45 expression are the immunophenotype abnormalities that often occur in abnormal B lymphoblasts, followed by the weakening of CD38 expression, the enhancement, weakening or loss of CD34 expression, the interruption or discontinuity of CD20 expression usually indicating the presence of abnormal cell groups, but still needs to be excluded from the therapeutic response after the application of CD20 monoclonal antibody; for B lymphoblastic leukemia with the original phenotype of CD10 positive, the enhancement of CD10 expression is a relatively easy immunophenotype abnormality, in addition, the leukemia cells will have the weakening of CD81 expression, the overexpression of CD58, CD73 and CD304, or the abnormal positive of CD66c and CD123.

[0032] In the present application, a 13-color one-tube detection scheme is successfully designed for the first time, which specifically includes designing a specific antibody combination, designing a fluorescein labeling scheme for the specific antibody combination, and further designing a flow analysis method in a targeted manner, including a specific gating method (quickly and efficiently obtaining and identifying cells), which is synergistic as a whole to ensure that the entire process is efficient and convenient, and further improve the detection rate, sensitivity, accuracy and detection density.

[0033] As a preferred technical solution, the detection unit in the device for detecting minimal residual disease is used to execute the following: staining the sample to be tested by using the antibody combination for detecting minimal residual disease in the first aspect, and performing flow cytometry detection by using a thirteen-color flow cytometer;

[0034] The analysis unit is used to execute the following:

[0035] The cells are gated, and the gating method includes:

[0036] Sequentially, the cell adhesion is removed by FSC-A and FSC-H double parameters; the live cell gate is selected by FSC-SSC; the blood cell gate is set by CD45-SSC sequential gating method; the CD19-SSC reverse gating method is used to analyze all B-lineage cells (including normal cells and abnormal cells); finally, the back gate is set, and the true and false of the cell population is judged by reversing the above gating sequence;

[0037] The CD19 gating method and the CD45 gating method are used, the CD19 gating method is used to analyze all B-lineage cells (including normal cells and abnormal cells) by gating, and the cell source is determined according to the antibody expression; the CD45 gating method is used to distinguish various types of cells in the sample, including lymphocytes, granulocytes, monocytes, blast cells and nucleated red blood cells.

[0038] Compared with the prior art, the present application has the following beneficial effects:

[0039] The antibody combination specifically used for detecting micro residual disease, combined with specific markers and gating strategies, realizes 13-color flow analysis, combined with composition and development process analysis of CD19+ cells and leukemia cell related immune phenotype (LAIP) analysis, can accurately lock the target cells, effectively distinguish normal cells and differential cells, significantly improve the detection rate and accuracy, and through controlling the number of obtained cells, the detection sensitivity is improved to 10 -5 . BRIEF DESCRIPTION OF DRAWINGS

[0040] Figure 1 The result graph of normal bone marrow CD19+ cell composition and development process;

[0041] Figure 2 The result graph of patient bone marrow, BCP-ALL patient sample MRD positive detection;

[0042] Figure 3 The result graph of patient bone marrow, BCP-ALL patient sample MRD positive detection;

[0043] Figure 4 The result graph of BCP ALL patient MRD positive detection. DETAILED DESCRIPTION

[0044] In order to further illustrate the technical means adopted by the present application and its effects, the present application will be further described below in conjunction with the embodiments and drawings. It can be understood that the specific embodiments described herein are only used to explain the present application, but not to limit the present application.

[0045] The specific technology or condition not mentioned in the examples is carried out according to the technology or condition described in the literature in the art, or according to the product instruction. The reagent or instrument not mentioned by the manufacturer is a conventional product that can be commercially available through a regular channel.

[0046] Example 1

[0047] The present embodiment provides a kit for detecting minimal residual disease, wherein the kit contains an antibody combination for detecting minimal residual disease, and the antibody combination comprises CD45 antibody, CD19 antibody, CD33 antibody, CD10 antibody, CD20 antibody, CD38 antibody, CD34 antibody, CD58 antibody, CD66c antibody, CD73 antibody, CD81 antibody, CD123 antibody and CD304 antibody, wherein each antibody in the antibody combination contains a fluorescein label, and the stock number and purchase manufacturer of each antibody are shown in Table 1 and Table 2, and the labeled fluorescein is in order as follows: V500, BV780, PECY7, APC-CY7, ECD, BV421, PercP-cy5.5, APC, FITC, PE, BV660, APC-A700 and BV610.

[0048] Table 1

[0049] Detection antibody CD33 CD19 CD10 CD20 CD34 CD38 CD45 Cat. No. A54824 302262 663486 IM3607U 343522 303526 56077 Supplier BC Biolend BD BC Biolend Biolend BD

[0050] Table 2

[0051] Detection antibody CD58 CD66c CD73 CD81 CD123 CD304 Cat. No. IM3701 IM2039U 550257 740590 B24028 354532 Supplier BC BC BD Biolend BC Biolend

[0052] Note: BD (Becton, Dickinson and Company, BD); BC (Beckman Coulter, BC).

[0053] The kit further comprises any one or a combination of at least two of the following: cell lysis solution, buffer or flow tube for use with a flow cytometer.

[0054] Embodiment 2

[0055] The present embodiment uses the kit in Embodiment 1 to process samples (derived from stem cell transplant donors).

[0056] The antibody combination for detecting minimal residual disease in Embodiment 1 is used to stain the sample to be tested, and the sample is detected on a three-laser thirteen-color flow cytometer of Beckman Coulter DxFLEX; the number of cells detected can be 2000000-5000000, including but not limited to 2500000, 3000000, 3500000, 4500000, 4600000, 4800000 or 4900000. The obtained cells are subjected to gate analysis, and the gate setting method comprises:

[0057] Sequentially remove cell adhesion by FSC-A and FSC-H double parameters; select live cell gate by FSC-SSC; set blood cell gate by CD45-SSC sequential gate setting method; set CD19-SSC reverse gate; set CD19-SSC reverse gate for B cell analysis (including normal cells and abnormal cells); finally, set back gate along the gate setting sequence in reverse, judge the true and false of the cell population, and quickly and accurately obtain the target analysis cells.

[0058] Suitable voltage and compensation are set through instrument quality control results and intracellular controls, and the number of cells required for the experiment is obtained accordingly. The expression profile based on the antibody combination distinguishes normal cells from abnormal cells.

[0059] According to the internationally recognized conservative value of 20 cells as the LOD (lowest detection limit), and taking the upper limit of the 95% confidence interval as the standard, LOD = 20 / number of cells obtained x 100%, and 50 cells are the minimum number of cells for repeatable quantitative detection generally accepted, LLOQ (lowest quantitative limit) = 50 / number of cells obtained x 100%, if 2,000,000 cells are obtained, then LOD and LLOQ are 0.001% and 0.003%, respectively, which is more sensitive than the internationally and domestically recognized MRD <0.01% for MRD negative, and its sensitivity can reach that of NGS gene sequencing.

[0060] The number of cells obtained is between 2,000,000 and 5,000,000, and based on the case where LOT is 20 events, the detection sensitivity is greater than or equal to 10 -5 (0.001%). Using the antibody combination scheme of the present application, the composition and development process of normal CD19+ cells can be clearly shown. Using the CD19 gate setting method to enclose all CD19 positive cell groups, it can be seen that the composition of normal cells includes CD19+CD20++ mature B lymphocytes (F gate), CD19+CD38++ plasma cells (E gate), CD10+CD38+CD34-CD20 sequential expression CD45 dim SSC small (A gate) and CD10+CD38+CD34+CD20-CD45dim SSC small (C gate) B progenitor cells, such as Figure 1 The composition and development process of normal bone marrow CD19+ cells are shown in the figure. The composition of normal cells includes CD19+CD20++ mature B lymphocytes (green), CD19+CD38++ plasma cells (dark purple), CD10+CD38+CD34-CD20 sequential expression CD45 dim SSC small (light orange) and CD10+CD38+CD34+CD20-CD45dim SSC small (dark orange) B progenitor cells.

[0061] The cell population appearing outside the relatively fixed positions can be defined as an abnormal cell population, and the enhancement of CD19 expression and the decrease of CD45 expression are often the immunophenotypic abnormalities of abnormal B lymphoblasts, followed by the decrease of CD38 expression, the enhancement, decrease or absence of CD34 expression, the interruption or discontinuity of CD20 expression, which usually indicates the presence of an abnormal cell population, but in specific analysis, the therapeutic response after the application of CD20 monoclonal antibody still needs to be excluded, and for B lymphoblastic leukemia with the original phenotype of CD10 positive, the enhancement of CD10 expression is a relatively easy immunophenotypic abnormality, in addition, the leukemia cells can exhibit the decrease of CD81 expression, the overexpression of CD58, CD73 and CD304, or the abnormal positivity of CD66c and CD123. Figure 2 As shown in the MRD positive detection result of a BCP ALL patient, using the analysis idea of DFN combined with LAIP, the normal mature lymphocyte population is shown in green, the normal B progenitor cell population is shown in yellow, a small amount of CD38++ plasma cells (dark purple), and an abnormal cell population with enhanced CD10 expression and absent CD38 expression appears outside the normally proliferating B progenitor cells, mature B lymphocytes and plasma cell population (Target gate), which does not express CD20 and CD45, and exhibits overexpression of CD73. Figure 3 As shown in the MRD positive detection result of a BCP ALL patient, using the analysis idea of DFN combined with LAIP, the normal mature lymphocyte population is shown in green, the normal B progenitor cell population is shown in yellow, a small amount of CD38++ plasma cells (dark purple), and the target cell population is shown in red, which is an abnormal B lymphocyte strongly expressing CD10 and CD58, expressing CD19 and CD66c, and not expressing CD20, CD38 and CD81.

[0062] Example 3

[0063] This example verifies the methodology of the detection scheme of the present application.

[0064] Take a bcr / abl(+) BCP-ALL sample, use the kit in Example 1 to perform flow detection and analysis on the sample, process the sample according to Example 2, and detect it on a Beckman Coulter DxFLEX three-laser thirteen-color flow cytometer, set appropriate voltages and compensations through instrument quality control results and cell population controls, and obtain the required cell number for the experiment.

[0065] The cell population appearing outside the relatively fixed positions can be defined as an abnormal cell population, and the enhancement of CD19 expression and the decrease of CD45 expression are often the immunophenotypic abnormalities of abnormal B lymphoblasts, followed by the decrease of CD38 expression, the enhancement, decrease or absence of CD34 expression, the interruption or discontinuity of CD20 expression, which usually indicates the presence of an abnormal cell population, but in specific analysis, the therapeutic response after the application of CD20 monoclonal antibody still needs to be excluded, and for B lymphoblastic leukemia with the original phenotype of CD10 positive, the enhancement of CD10 expression is a relatively easy immunophenotypic abnormality, in addition, the leukemia cells can exhibit the decrease of CD81 expression, the overexpression of CD58, CD73 and CD304, or the abnormal positivity of CD66c and CD123. Figure 4The MRD positive detection result of BCP ALL patients is shown in the figure. The analysis idea of DFN combined with LAIP is used. The normal mature lymphocyte group is shown in green, the normal B progenitor cell group is shown in yellow, a small amount of CD38++ plasma cells (dark purple), and the target cell group is shown in red. In addition to the normal proliferating B progenitor cells and mature B lymphocytes and plasma cell groups, an abnormal cell group (shown in red) with enhanced CD10 expression and absent CD38 expression appears. This abnormal cell group also has abnormal expression of CD123. It is indicated that there are leukemia cells in the sample, which is consistent with the bcr / abl(+) result of molecular biology, and the accuracy meets the standard. The number of cells obtained in the experiment is between 2,000,000 and 5,000,000. Based on the LOT of 20 Events, the detection sensitivity is greater than or equal to 10 -5 (0.001%). Five flow cytometry detections were performed on the sample, and the results were all positive. The detection values (the proportion of MRD positive cells in the total number of cells obtained) were 1.562%, 1.564%, 1.560%, 1.561%, and 1.562%, respectively. The precision of this detection is 0.09.

[0066] Example 4

[0067] In this example, 119 MRD positive samples were detected by the method of Example 2. According to the LAIP phenotype combined with the DFN abnormal cell group, the detection value > 0.01% was used as the MRD positive judgment standard. 114 MRD samples were detected, and the detection rate (detection rate = the number of detected MRD samples / 119*100%) was 95.8%. This indicates that the method of the present application can efficiently detect MRD residues.

[0068] Comparative Example 1

[0069] In this comparative example, 119 MRD positive samples in Example 4 were detected by the commonly used 8-color flow detection method. The antibodies used were: CD58-FITC, CD10-APC-CY7, CD19-APC, CD34-PerCP-Cy5.5, CD20-APC-CY7, CD38-PE-CY7, CD123-BV421 and CD45-V500. According to the LAIP phenotype combined with the DFN abnormal cell group, the detection value > 0.01% was used as the MRD positive judgment standard. 103 MRD samples were detected, and the detection rate was 86.5%. This indicates that the method of the present application can further improve the MRD detection rate compared with the existing detection method.

[0070] Comparative Example 2

[0071] In this comparative example, flow detection was performed, and the only difference compared with Example 4 was that the CD33 antibody was replaced with an equal amount of CD65 antibody. 105 samples were detected, and the detection rate was 88.2%.

[0072] Comparative Example 3

[0073] The present comparative example was subjected to flow detection, and compared with Example 4, only CD66c antibody was replaced by CD15 antibody. 109 cases were detected, and the detection rate was 91.5%.

[0074] Comparative Example 4

[0075] The present comparative example was subjected to flow detection, and compared with Example 4, only CD73 antibody was replaced by CD2 antibody. 107 cases were detected, and the detection rate was 89.9%.

[0076] Comparative Example 5

[0077] The present comparative example was subjected to flow detection, and compared with Example 4, only CD81 antibody was replaced by CD56 antibody. 110 cases were detected, and the detection rate was 92.4%.

[0078] Comparative Example 6

[0079] The present comparative example was subjected to flow detection, and compared with Example 4, only CD304 antibody was replaced by CD7 antibody. 101 cases were detected, and the detection rate was 84.8%.

[0080] It can be seen that the overall detection rate of Comparative Examples 2-6 is significantly lower than that of Example 4, indicating that the specific antibody marker combination designed in the present application can further improve the detection rate.

[0081] In summary, the specific antibody combination for detecting minimal residual disease designed in the present application, the specific flow detection process, the realization of 13-color flow analysis, the analysis of the composition and development process of CD19+ cells and the analysis of leukemia cell-related immunophenotype (LAIP), can accurately lock the target cells, effectively distinguish normal cells and differential cells, significantly improve the detection rate and accuracy, and by controlling the number of cells obtained, the detection sensitivity is improved to 10 -5 .

[0082] The applicant declares that the present application is illustrated by the above examples, but the present application is not limited to the above detailed method, that is, it does not mean that the present application must rely on the above detailed method to be implemented. It should be understood by those skilled in the art that any improvement of the present application, equivalent replacement of each raw material of the product of the present application, addition of auxiliary ingredients, selection of specific methods, etc. fall within the protection scope and disclosure scope of the present application.

Claims

1. An antibody combination for detecting minimal residual disease, characterized in that, The antibody combination comprises CD45 antibody, CD19 antibody, CD33 antibody, CD10 antibody, CD20 antibody, CD38 antibody, CD34 antibody, CD58 antibody, CD66c antibody, CD73 antibody, CD81 antibody, CD123 antibody and CD304 antibody.

2. The antibody combination for detecting minimal residual disease according to claim 1, characterized by, The antibodies in the antibody combination all contain fluorescein label.

3. The antibody combination for detecting minimal residual disease according to claim 1 or 2, characterized in that, The CD45 antibody, CD19 antibody, CD33 antibody, CD10 antibody, CD20 antibody, CD38 antibody, CD34 antibody, CD58 antibody, CD66c antibody, CD73 antibody, CD81 antibody, CD123 antibody and CD304 antibody are sequentially labeled with V500, BV780, PECY7, APC-CY7, ECD, BV421, PercP-cy5.5, APC, FITC, PE, BV660, APC-A700 and BV610.

4. The antibody combination for use in detecting minimal residual disease according to any one of claims 1 to 3, characterized in that, The antibodies in the antibody combination are all monoclonal antibodies.

5. Use of the antibody combination for detecting minimal residual disease according to any one of claims 1-4 in the manufacture of a product for detecting minimal residual disease.

6. A kit for detecting minimal residual disease, characterized in that, The kit comprises the antibody combination for detecting minimal residual disease according to any one of claims 1-4; Preferably, the kit further comprises any one or a combination of at least two of cell lysate, buffer or flow tube for use with flow cytometer.

7. A device for detecting minimal residual disease, characterized in that The device comprises a detection unit and an analysis unit; The detection unit is configured to perform operations comprising: staining the sample to be tested with the antibody combination for detecting minimal residual disease according to any one of claims 1-4, and performing flow cytometry detection; The analysis unit is configured to perform operations comprising: gating the cells, and distinguishing normal cells and abnormal cells based on the expression profile of the antibody combination.

8. The device for detecting minimal residual disease according to claim 7, characterized in that, The instrument for flow cytometry detection comprises a thirteen-color flow cytometer.

9. The device for detecting minimal residual disease according to claim 7, characterized in that, The gating method comprises: using FSC-A and FSC-H double parameters to remove cell adhesion; using FSC-SSC to select live cell gate; using CD45-SSC sequential gating method to set blood cell gate, using CD19-SSC reverse gating method to circle gate all B-lineage cells for analysis; finally, back gate setting, along the gating sequence in reverse, to judge the true or false of the cell population; using CD19 gating method and CD45 gating method, the CD19 gating method is used to circle gate all B-lineage cells containing normal cells and abnormal cells for analysis, and the cell source is determined according to the antibody expression; the CD45 gating method is used to distinguish various types of cells in the sample, including lymphocytes, granulocytes, monocytes, blast cells and nucleated red blood cells.

10. The device for detecting minimal residual disease according to any one of claims 7-9, wherein The detection unit is configured to perform operations comprising: staining the sample to be tested with the antibody combination for detecting minimal residual disease according to any one of claims 1-4, and performing flow cytometry detection with a thirteen-color flow cytometer; The analysis unit is configured to perform operations comprising: gating the cells, and the gating method comprises: The FSC-A and FSC-H double parameters are used to remove cell adhesion; the FSC-SSC is used to select a live cell gate; the CD45-SSC sequential gate setting method is used to set a blood cell gate; the CD19-SSC reverse gate setting method is used to circle gate all B-lineage cells for analysis; finally, the gate setting is set, and the cell population is judged to be true or false along the gate setting sequence in reverse. The CD19 gate setting method is used to circle gate all B-lineage cells containing normal cells and abnormal cells for analysis, and the cell source is determined according to the antibody expression; the CD45 gate setting method is used to distinguish various types of cells in the sample, including lymphocytes, granulocytes, monocytes, blast cells and nucleated red blood cells.

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