CN117192121A

CN117192121A - Antibody composition for MDS and/or AML minimal residual disease detection and application thereof

Info

Publication number: CN117192121A
Application number: CN202311103981.8A
Authority: CN
Inventors: 黄晓军; 常英军; 赵晓甦; 李思琦; 吴蔚冰
Original assignee: Peking University Peoples Hospital
Current assignee: Peking University Peoples Hospital
Priority date: 2023-08-30
Filing date: 2023-08-30
Publication date: 2023-12-08

Abstract

The application discloses an antibody composition for detecting MDS and/or AML minimal residual disease and application thereof. Specifically disclosed are compositions comprising CD45RA antibodies, CLL-1 antibodies, TIM-3 antibodies, CD7 antibodies, CD11b antibodies, CD22 antibodies, CD56 antibodies, CD123 antibodies, CD33 antibodies, CD38 antibodies, CD44 antibodies, CD34 antibodies, CD45 antibodies, HLA-DR antibodies, and CD117 antibodies. The application also discloses a fluorescent label corresponding to the antibody, an MRD detection method and a result interpretation method. The application can more sensitively and earlier detect and identify residual cells of MDS and/or AML by only using 15 antibody compositions and one tube of cells for one-time sampling, evaluate the curative effect of treatment, reduce the requirement on sample quantity and the operation quantity and improve the sensitivity to the detection of MDS and/or AML tiny residual diseases.

Description

Antibody composition for MDS and/or AML minimal residual disease detection and application thereof

Technical Field

The application belongs to the field of biological medicine, and in particular relates to an antibody composition for detecting myelodysplastic syndrome (Myelodysplastic syndrome, MDS) and/or acute myelogenous leukemia (Acute Myeloid Leukemia, AML) minimal residual disease and application thereof.

Background

Myelodysplastic syndrome (MDS) and Acute Myelogenous Leukemia (AML) are serious diseases that seriously jeopardize human health; recurrence is the primary cause of treatment failure, and minimal residual disease (Minimal residual disease, MRD) is the root cause of recurrence. Currently, MRD detection methods for MDS and AML include Multiparameter Flow Cytometry (MFC), real-time quantitative polymerase chain reaction (RT-PCR), and second generation sequencing techniques (NGS). MFC is a detection technique that uses a combination of multiple antibodies labeled with different fluorescent markers to detect the expression status of hematopoietic cell surface or intracellular antigens, and further analyzes and determines whether the cells are abnormal in serial sources, differentiation degrees, and phenotypes. MFC can cover more than 90% of the population, and is the most commonly used MRD detection method for MDS and AML. However, the traditional MFC detection MRD method is based on Leukemia Associated Immunophenotype (LAIP) and differentiation (D-f-N) from normal myeloid phenotype, and the internationally accepted threshold for MRD detection is 0.1% (low sensitivity), with high false negative, false positive problems. In particular, for MDS patients, the presence of pathological hematopoiesis in this class of patients has a serious impact on the traditional detection of residual disease in MFC. Therefore, it is necessary to establish new MFC antibody combinations to detect MDS and AML residual disease to increase the sensitivity of detection, reduce the false negative rate and false positive rate.

Disclosure of Invention

The technical problem to be solved by the application is how to more sensitively and more specifically detect MDS and/or AML minimal residual disease; the technical problems to be solved are not limited to the described technical subject matter, and other technical subject matter not mentioned herein will be clearly understood by those skilled in the art from the following description.

To solve the above technical problems, the present application provides an antibody composition for detecting MDS and/or AML minimal residual disease, which comprises CD45RA antibody, CLL-1 antibody, TIM-3 antibody, CD7 antibody, CD11b antibody, CD22 antibody, CD56 antibody, CD123 antibody, CD33 antibody, CD38 antibody, CD44 antibody, CD34 antibody, CD45 antibody, HLA-DR antibody and CD117 antibody.

Further, each antibody in the antibody composition may be a monoclonal antibody.

Further, each antibody in the antibody composition may be a fluorescein-labeled antibody.

Further, the fluorescein labels of the CD45RA antibody, the CLL-1 antibody, the TIM-3 antibody, the CD7 antibody, the CD11b antibody, the CD22 antibody, the CD56 antibody, the CD123 antibody, the CD33 antibody, the CD38 antibody, the CD44 antibody, the CD34 antibody, the CD45 antibody, the HLA-DR antibody and the CD117 antibody can be as follows: FITC, PE, PE, PE, PE, PE, PE, perCP-Cy5.5, PE-Cy7, APC-H7, BV421, V500, alexa 700 and BV605.

The application also provides a kit for the detection of MDS and/or AML minimal residual disease, which may comprise an antibody composition as described herein.

Further, the kit may further comprise a red blood cell lysate, brilliant Stain Buffer and/or a buffer.

The buffer may be a PBS buffer.

The application also provides any one of the following uses of the antibody composition described herein:

a1 Use of the composition for the preparation of a product for the detection of MDS and/or AML minimal residual disease;

a2 For the preparation of a product for detecting residual tumor cells in an ex vivo sample during or after MDS treatment;

a3 Use of the composition for the preparation of a product for detecting residual tumor cells in an ex vivo sample during or after AML treatment;

a4 For the preparation of a product for the evaluation of the efficacy of MDS and/or AML.

The efficacy assessment may be based on the level of MRD (i.e., the percentage of residual tumor cells in bone marrow or peripheral blood nucleated cells).

The application also provides a system for detection of minimal residual disease in MDS and/or AML, the system may include a detection portion and an analysis portion, wherein:

the detection part can comprise a reagent for detecting MDS or AML cells in a sample to be detected by multi-parameter flow cytometry, so as to obtain a detection result of the sample to be detected; the agent comprises an antibody composition as described herein; the antibody composition was used in 1 flow tube at the time of detection;

the assay portion can be used to analyze the MDS or AML cells detected by the detection portion, determine the phenotype, number (proportion of nucleated cells) of the cells, and determine whether residual MDS or AML cells are present in the test sample.

Further, the system, when used to detect MDS or AML cells in a test sample, may comprise the steps of:

preparing a flow on-cell machine sample after treatment of a sample to be tested with an antibody composition or the kit as described herein; performing flow cell on-machine detection; wherein the flow cell is provided with a gate during the on-machine detection according to the following mode:

setting a R1 living cell gate, removing fragments and dead cells, and setting a lymphocyte gate (R2), a monocyte gate (R3), a granulocyte gate (R4), a nucleated erythrocyte gate (R5) and a naive cell gate (R6) in the R1 gate by using CD 45/SSC; and analyzing the expression condition of the antigens in different cell doors.

Further, the analysis of the expression of antigens in different cell gates may include any of the following:

b1 Using SSC/CD34 dot patterns to distinguish CD34 in R6 according to the expression or non-expression of CD34 ⁺ Cells (R7), and CD34 in R7 was discriminated from each other by using a CD34/CD38 dot pattern according to the presence or absence of CD38 expression ⁺ CD38 ^- Cells showing the expression ratios of CD33, CD123, CD45RA, CD44, CD117, HLA-DR and the mixed antibodies cocktail (CD 7, CD11b, CD22, CD56, CLL-1 and TIM-3) on CD34+CD38-cells, CD34 was calculated ⁺ CD38 ^- CD33 ⁺ Cells, CD34 ⁺ CD38 ^- CD123 ⁺ Cells, CD34 ⁺ CD38 ^- CD45RA ⁺ Cells, CD34 ⁺ CD38 ^- CD44 ⁺ Cells, CD34 ⁺ CD38 ^- CD117 ⁺ Cells, CD34 ⁺ CD38 ^- HLA-DR ⁺ Cells and CD34 ⁺ CD38 ^- cocktail ⁺ The proportion of cells occupying the nucleus;

b2 Using CD45/CD117 spot diagram to distinguish CD117 in R1 according to CD117 expression or not ⁺ Cell (R8), benefitingBy using a CD117/CD34 dot diagram, CD117+CD34-cells in R8 are distinguished according to the expression or non-expression of CD34, and CD117 is displayed ⁺ CD34 ^- The expression ratios of CD33, CD123, CD45RA, CD44, CD117, HLA-DR and cocktail on the cells were calculated to calculate CD117 ⁺ CD34 ^- CD33 ⁺ Cell, CD117 ⁺ CD34 ^- CD123 ⁺ Cell, CD117 ⁺ CD34 ^- CD45RA ⁺ Cell, CD117 ⁺ CD34 ^- CD44 ⁺ Cell, CD117 ⁺ CD34 ^- CD117 ⁺ Cell, CD117 ⁺ CD34 ^- HLA-DR ⁺ Cells and CD117 ⁺ CD34 ^- cocktail ⁺ The cells occupy a proportion of nuclear cells.

The sample to be tested is an in-vitro sample, and can be a bone marrow sample, a peripheral blood sample, hydrothorax, ascites or cerebrospinal fluid.

The purpose of the use described herein may be a disease diagnosis purpose, a disease prognosis purpose and/or a disease treatment purpose, as well as a non-disease diagnosis purpose, a non-disease prognosis purpose and a non-disease treatment purpose; their direct purpose may be information of intermediate results of obtaining disease diagnosis results, disease prognosis results and/or disease treatment results, and their direct purpose may be non-disease diagnosis purpose, non-disease prognosis purpose and/or non-disease treatment purpose.

Aiming at the problems existing in the traditional multi-parameter flow cytometry (MFC) detection of MDS and AML: (1) the detection sensitivity is low, and the internationally accepted threshold value is only 0.1%; (2) the problems of high false negative and false positive exist; (3) the present application devised a set of 15 antibody compositions for MDS and/or AML residual cell detection by MFC technology to solve the above problems.

Through extensive and intensive research, screening, optimizing antibody combinations, fluorescent label combinations of corresponding antibodies and innovative result interpretation methods, the application can detect and identify residual cells of MDS and/or AML more sensitively and earlier by only using one tube of 15 antibody compositions and one tube of cells for one-time sample application, evaluate treatment efficacy and predict disease recurrence.

Experiments prove that compared with the prior art, the application has the following beneficial effects:

at present, 8-10 color antibody combinations are generally used for carrying out traditional MFC residual disease detection on MDS or AML, each disease needs to detect 2-3 tube antibody combinations (18-22 antibodies), but the application only uses 1 tube antibody combination (15 antibodies) to carry out residual disease detection on MDS and/or AML, so that the requirements on sample quantity and operation quantity are reduced, the labor intensity is lightened, the operation time is saved, the repeated application of door setting antibodies is reduced, the use quantity of effective antibodies is increased, whether 15 antibodies are simultaneously expressed or not can be simultaneously observed, the phenotype relation of the mutual combination of the antibodies is analyzed, and the sensitivity on the detection on MDS and/or AML tiny residual disease is improved. In addition, the application also discloses the identification of CD34 by using two-dimensional maps such as CD117/cocktail and the like ^- An analytical method for minimal residual disease in MDS or AML patients, which is used for detecting the minimal residual disease in patients which do not express CD34 antigen on MDS or AML stem progenitor cells. The antibody composition of the application has very high specificity and sensitivity for detecting myelodysplastic syndrome (MDS) and/or Acute Myelogenous Leukemia (AML) tiny residual disease, the sensitivity can reach 90%, and the application has wide clinical application value.

Drawings

FIG. 1 shows an LSC gating analysis method. The sample to be tested is a normal bone marrow.

FIG. 2 shows an example of LSC detection results for AML patients.

FIG. 3 shows an example of LSC detection results in patients with MDS.

FIG. 4 is CD117 ⁺ CD34 ^- Cell gating assay. The specimen is a normal bone marrow.

FIG. 5 shows an example of CD117 of AML primary patients ⁺ CD34 ^- Cell detection results.

FIG. 6 is an example of CD117 of a patient with MDS ⁺ CD34 ^- Cell detection results.

Detailed Description

The following detailed description of the application is provided in connection with the accompanying drawings that are presented to illustrate the application and not to limit the scope thereof. The examples provided below are intended as guidelines for further modifications by one of ordinary skill in the art and are not to be construed as limiting the application in any way.

The experimental methods in the following examples, unless otherwise specified, are conventional methods, and are carried out according to techniques or conditions described in the literature in the field or according to the product specifications. Materials, reagents and the like used in the examples described below are commercially available unless otherwise specified.

The following examples employ flow cytometry to analyze and detect Minimal Residual Disease (MRD) in bone marrow fluid specimens of clinical patients. The bone marrow fluid specimen includes: 20 healthy donor bone marrow samples, 123 primary AML patient bone marrow samples, 31 post-chemotherapy AML patient bone marrow samples, 100 pre-transplant, post-transplant follow-up bone marrow samples. The samples are from Beijing university people's hospital, and the sample collection is approved by the ethical committee of the Beijing university people's hospital, approval number: 2023PHB164-001.

Brilliant Stain Buffer in the following examples is BD company product, cat No.: 563794.

antibodies in the following examples are commercially available as they are, and antibodies according to examples of the present application are purchased from BD company, biolegend company.

The luciferin in the following examples includes: FITC (Biolegend, cat# 304106), PE (Biolegend, cat# 343106, 301306, 363504, 345006 and BD, cat# 556647, 562566), perCP-Cy5.5 (Biolegend, cat# 306016), PE-Cy7 (Biolegend, cat# 303434), APC (Biolegend, cat# 303510), APC-H7 (Biolegend, cat# 103028), BV421 (Biolegend, cat# 343610), V500 (BD, cat# 560777), alexa 700 (BD, cat# 560743) and BV605 (BD, cat# 562687).

Example 1 antibody compositions for detection of MDS and/or AML minimal residual disease

Minimal Residual Disease (MRD) refers to the presence of malignant cells that remain in the body of a cancer patient during or after treatment. MRD causes postoperative tumor recurrence in cancer patients, so cancer patients need to monitor MRD, monitoring postoperative tumor recurrence risk. MRD detection may refer to the detection of microscopic or invisible malignant cells remaining in the body during or after treatment.

The present example provides an antibody composition for MDS and/or AML minimal residual disease detection based on Multiparameter Flow Cytometry (MFC), 15 antibodies 10-color fluorescein, antibody names and fluorescent markers are shown in Table 1. Brilliant Stain Buffer was mixed with 15 antibodies in table 1 in 1 container for determination of minimal residual disease in MDS and AML specimens.

The preparation method of the corresponding fluorescein-labeled monoclonal antibody premix in Table 1 is as follows: brilliant Stain Buffer 20. Mu.l/person was first added to 1 vessel, and the surface antibody was added in the amounts shown in Table 1 and mixed well.

The antibody composition is prepared into a detection kit for detecting MDS and/or AML sample tiny residual diseases. The kit also comprises a red blood cell lysate and PBS, wherein the red blood cell lysate can be prepared by self or can be purchased commercially (such as BD company product, product number 349202).

TABLE 1 names, fluorescent markers and amounts of 15 antibodies of the application

Antibody name	Fluorescein (Lu)	Dosage of	Antibody name	Fluorescein (Lu)	Dosage of
						CD45RA	FITC	2.5 μl/person times	CD33	PE-Cy7	5.0 μl/person times
CLL-1	PE	2.5 μl/person times	CD38	APC	1.0 μl/person times
						TIM-3	PE	2.5 μl/person times	CD44	APC-H7	2.5 μl/person times
CD7	PE	2.5 μl/person times	CD34	BV421	2.5 μl/person times
						CD11b	PE	2.5 μl/person times	CD45	V500	3.0 μl/person times
CD22	PE	2.5 μl/person times	HLA-DR	Alexa 700	2.5 μl/person times
						CD56	PE	10.0 μl/person times	CD117	BV605	2.5 μl/person times
CD123	PerCP-Cy5.5	2.5 μl/person times

Example 2 use of antibody compositions for detection of MDS and/or AML minimal residual disease

The 15 antibody compositions of the application are used for flow cytometry to analyze the minimal residual disease of MDS and/or AML patients, and the specific method is as follows:

1. sample collection

1-3mL of the obtained human bone marrow fluid is immediately placed in a heparin anticoagulation tube and is rapidly reversed for several times to prevent the sample from solidifying, and the sample is sent to a laboratory as soon as possible after being collected and is placed at 4 ℃ for cold storage. MFC testing must be completed within 48 hours.

2. Sample preparation and detection

(1) Cell count: taking 10 mu L of bone marrow, adding 150 mu L of PBS, mixing well, and counting each microliter of fine powderCell number, cell concentration was adjusted to 5X 10 based on the detection result ⁶ ～10×10 ⁶ mu.L/100. Mu.L of cells were taken and added to the flow tube in an amount of 50. Mu.L to 100. Mu.L.

(2) Antigen staining:

a) Washing: 1mL of a flow tube containing 0.1% NaN was added to the flow tube of step (1) ₃ And 1% -2% BSA PBS wash, 300g centrifugal washing 5min, discarding the supernatant, washing 2 times. Each tube was added with a premix (61.5. Mu.l) of the corresponding fluorescein-labeled monoclonal antibody of Table 1, and incubated at room temperature for 15min in the absence of light.

b) Hemolysis: 2mL of 1 XFACS hemolysin is added, the mixture is uniformly mixed by low-speed vortex, and the mixture is kept stand for 8 to 10 minutes at room temperature in a dark place. Centrifuge wash at 300g for 5min and discard supernatant.

c) Washing: 1mL of a solution containing 0.1% NaN was added ₃ And 1% -2% BSA PBS wash, 300g centrifugal washing 5min, discard the supernatant. 200. Mu.L of PBS was added to the suspension and the suspension was checked on-line.

(3) And (3) detecting:

a) Determining an optimum voltage and compensation: the voltage is set according to the conventional operation method of the spectrum flow cytometer, and a single-dye sample is prepared by referring to the fluorescent color matching of the kit for instrument setting.

b) Instrument setup, calibration and quality control: BDFACS Cantoplus starts the preheating machine for more than 20min and is rinsed by deionized water, and the inner quality control product is detected, so that each detection value is ensured to be within a control range. And (5) calling PANAL for loading and collecting data.

c) And (3) detecting: according to the set instrument conditions, 100 ten thousand cells are obtained per tube. If the detection can not be carried out on the machine in time, 0.5mL of 1% paraformaldehyde is added, and the mixture is uniformly mixed and then is stored in a refrigerator at 4 ℃ for 24 hours to finish the detection.

3. Data analysis

Data were analyzed using Kaluza software as follows:

(1) The FSC/SSC two-dimensional map was used to remove debris, adherent cells, and dead cells, and viable single cells were gated on R1.

(2) R1 portal cells are displayed, a CD45/SSC two-dimensional map is established, and lymphocytes, monocytes, granulocytes, nucleated erythrocytes and naive cells are gated and set with different colors according to the difference of the distribution of the CD45 and the SSC. Lymphocytes (R2): CD45 highest/SSC lowest; monocytes (R3): CD45 is lower than lymphocytes/SSC is higher than lymphocytes and lower than granulocytes; granulocytes (R4): CD45 is lower than monocytes/SSC maximum; nucleated red blood cells (R5): CD45 negative/SSC is lower than lymphocytes or the same; naive cells (R6): CD45 is lower/SSC than lymphocytes and higher or the same. In normal bone marrow, each population of cells has a normal range of proportions: 20% -40% of lymphocytes, 2% -8% of monocytes, 40% -60% of granulocytes, 2% -15% of nucleated erythrocytes and less than 5% of naive cells. And observing whether the proportion of each group of cells is normal, whether the proportion of the naive cells is increased, and the like.

(3) The expression of the antigen of the application in naive cells was mainly analyzed. The expression of the antigen on normal blood cells is shown in Table 2. By using SSC/CD34 dot pattern, based on CD34 expression or not, the CD34 in R6 is distinguished ⁺ Cells (R7). By using the CD34/CD38 dot pattern, the CD34 in R7 is distinguished according to whether the CD38 is expressed or not ⁺ CD38 ^- Cells, display CD34 ⁺ CD38 ^- CD34 was calculated by calculating the expression ratios of CD33, CD123, CD45RA, CD44, CD117, HLA-DR and cocktail (CD 7, CD11b, CD22, CD56, CLL-1 and TIM-3) on the cells ⁺ CD38 ^- CD33 ⁺ Cells, CD34 ⁺ CD38 ^- CD123 ⁺ Cells, CD34 ⁺ CD38 ^- CD45RA ⁺ Cells, CD34 ⁺ CD38 ^- CD44 ⁺ Cells, CD34 ⁺ CD38 ^- CD117 ⁺ Cells, CD34 ⁺ CD38 ^- HLA-DR ⁺ Cells and CD34 ⁺ CD38 ^- cocktail ⁺ The cells occupy a proportion of nuclear cells. Normal Hematopoietic Stem Cell (HSC) phenotype is CD34 ⁺ CD38 ^- But did not express CD33, CD123, CD45RA and the mixed antibodies cocktail (CD 7, CD11b, CD22, CD56, CLL-1 and TIM-3), CD44 expression was not enhanced. The present application thus utilizes the above antibodies to identify HSCs and LSCs (leukemia stem cells) or leukemia stem cell-like MDS cells.

TABLE 2 expression of antigens on Normal cells

Some MDS or AML specimens have residual disease cells that do not express CD34, and thus the present application uses a CD117/CD34 two-dimensional dot pattern to analyze CD117 ⁺ CD34 ^- And (3) cells. By using a CD45/CD117 dot pattern, the CD117 in R1 is distinguished according to whether the CD117 is expressed or not ⁺ Cells (R8). By using the CD117/CD34 dot pattern, the CD117 in R8 is distinguished according to whether the CD34 is expressed or not ⁺ CD34 ^- Cells, displaying CD117 ⁺ CD34 ^- The expression ratios of CD33, CD123, CD45RA, CD44, CD117, HLA-DR and cocktail on the cells were calculated to calculate CD117 ⁺ CD34 ^- CD33 ⁺ Cell, CD117 ⁺ CD34 ^- CD123 ⁺ Cell, CD117 ⁺ CD34 ^- CD45RA ⁺ Cell, CD117 ⁺ CD34 ^- CD44 ⁺ Cell, CD117 ⁺ CD34 ^- CD117 ⁺ Cell, CD117 ⁺ CD34 ^- HLA-DR ⁺ Cells and CD117 ⁺ CD34 ^- cocktail ⁺ The cells occupy a proportion of nuclear cells.

4. Experimental results

(1) A total of 20 healthy donor bone marrow samples were tested using the antibody compositions of the present application. FIG. 1 shows that HSCs in normal bone marrow of healthy donors very low expressed CD33, CD123, CD45RA and the mixed antibody cocktail (CD 7, CD11b, CD22, CD56, CLL-1 and TIM-3), normally expressing CD44. FIG. 4 shows CD117 in normal bone marrow of healthy donors ⁺ CD34 ^- Cellular antibody expression.

(2) Using the antibody composition of the present application, a total of 123 bone marrow samples from primary AML patients and 31 bone marrow samples from AML patients after chemotherapy were examined. The samples were also subjected to conventional 8-color 1-5 tube antibody combinatorial immunophenotyping or minimal residual disease detection.

FIG. 2 shows an example of LSC detection results of AML primary patients. The ratio of naive (R6) nucleated cells from CD45/SSC was 58.7%, CD34 in the R6 gate ⁺ The proportion of cells (R7) occupying the nucleus was 6.7%, in which part of CD34+CD38-cells abnormally expressed CD33CD45RA, CD123, CD117, HLA-DR, cocktail, and LSC are present. The detection result is consistent with the conventional detection result by using the method. FIG. 5 shows one example of CD117 of AML primary patients ⁺ CD34 ^- Cell detection results. Display of CD117 from CD45/CD117 ⁺ CD34 ^- The proportion of cells (R8) occupied the nucleated cells was 27.9%, and some of them showed enhanced CD44 expression, increased CD45RA expression, CD123, CD33, cocktail, HLA-DR expression, and abnormal cells were present. The detection result is consistent with the conventional detection result by using the method.

(3) Using the antibody compositions of the application, a total of 100 MDS patients were examined for pre-transplant, post-transplant follow-up bone marrow samples. The samples were also subjected to conventional 8-color 1-2 tube (Wang XR et al, ann Hematol 2020;99 (2): 215-221.) antibody combination minimal residual disease detection.

The results showed that 10 patients who were positive for minimal residual disease were relapsed using the antibody composition of the present application, of which only 6 conventional MFCs were positive for minimal residual disease, whereas 9 patients who were positive for minimal residual disease were detected using the antibody composition of the present application. The above results demonstrate that the antibody combinations of the application are highly sensitive (90% vs. 60%) to detection and recurrence prediction of minimal residual disease in MDS compared to conventional MFCs.

FIG. 3 shows one example of LSC detection results prior to transplantation in MDS patients. The ratio of naive (R6) nucleated cells was 3.96% from CD45/SSC, CD34 in the R6 gate ⁺ The proportion of cells (R7) occupying the nucleus was 1.06%, part of which was CD34 ⁺ CD38 ^- Cells abnormally expressed CD33, CD45RA, CD123, CD117, HLA-DR, cocktail, and LSC was present. The detection result is consistent with the conventional detection result by using the method. FIG. 6 shows one example of CD117 prior to transplantation in MDS patients ⁺ CD34 ^- Cell detection results. Display of CD117 from CD45/CD117 ⁺ CD34 ^- The proportion of cells (R8) occupied the nuclear cells was 1.2%, and some of them expressed CD45RA and CD33 were increased, and abnormal cells were present. The detection result is consistent with the conventional detection result by using the method.

The present application is described in detail above. It will be apparent to those skilled in the art that the present application can be practiced in a wide range of equivalent parameters, concentrations, and conditions without departing from the spirit and scope of the application and without undue experimentation. While the application has been described with respect to specific embodiments, it will be appreciated that the application may be further modified. In general, this application is intended to cover any variations, uses, or adaptations of the application following, in general, the principles of the application and including such departures from the present disclosure as come within known or customary practice within the art to which the application pertains.

Claims

1. An antibody composition for use in the detection of MDS and/or AML minimal residual disease, comprising a CD45RA antibody, CLL-1 antibody, TIM-3 antibody, CD7 antibody, CD11b antibody, CD22 antibody, CD56 antibody, CD123 antibody, CD33 antibody, CD38 antibody, CD44 antibody, CD34 antibody, CD45 antibody, HLA-DR antibody, and CD117 antibody.

2. The antibody composition of claim 1, wherein each antibody in the antibody composition is a monoclonal antibody.

3. The antibody composition of claim 1 or 2, wherein each antibody in the antibody composition is a fluorescein-labeled antibody.

4. The antibody composition of any one of claims 1-3, wherein the fluorescein labels of the CD45RA antibody, CLL-1 antibody, TIM-3 antibody, CD7 antibody, CD11b antibody, CD22 antibody, CD56 antibody, CD123 antibody, CD33 antibody, CD38 antibody, CD44 antibody, CD34 antibody, CD45 antibody, HLA-DR antibody, and CD117 antibody are, in order: FITC, PE, PE, PE, PE, PE, PE, perCP-Cy5.5, PE-Cy7, APC-H7, BV421, V500, alexa 700 and BV605.

5. A kit for the detection of MDS and/or AML minimal residual disease, characterized in that the kit comprises an antibody composition according to any one of claims 1-4.

6. The kit of claim 5, further comprising a red blood cell lysate, brilliant Stain Buffer, and/or a buffer.

7. Use of the antibody composition of any one of claims 1-4 for any one of the following:

a3 Use of a product for detecting residual tumor cells in an ex vivo sample during or after AML treatment;

8. A system for the detection of micro residual disease in MDS and/or AML, the system comprising a detection portion and an analysis portion, wherein:

the detection part comprises a reagent for detecting MDS or AML cells in a sample to be detected by multi-parameter flow cytometry, and a detection result of the sample to be detected is obtained; the agent comprises the antibody composition of any one of claims 1-4; the antibody composition was used in 1 flow tube at the time of detection;

the analysis part is used for analyzing the MDS or AML cells detected by the detection part, determining the phenotype and the quantity of the cells, and determining whether residual MDS or AML cells exist in the sample to be detected.

9. The system of claim 8, wherein the system, when used to detect MDS or AML cells in a test sample, comprises the steps of:

preparing a flow cytometric sample after treatment of a sample to be tested with the antibody composition of any one of claims 1-4 or the kit of claim 5 or 6; performing flow cell on-machine detection; wherein the flow cell is provided with a gate during the on-machine detection according to the following mode:

setting a R1 living cell gate, removing fragments and dead cells, and setting a lymphocyte gate, a monocyte gate, a granulocyte gate, a nucleated erythrocyte gate and a naive cell gate in the R1 gate by using CD 45/SSC; and analyzing the expression condition of the antigens in different cell doors.

10. The system of claim 9, wherein said analyzing the expression of antigens in different cell gates comprises any of the following:

b1 Using SSC/CD34 dot patterns to distinguish CD34 in R6 according to the expression or non-expression of CD34 ⁺ Cells, using a CD34/CD38 dot map, distinguish CD34 in R7 based on whether CD38 is expressed or not ⁺ CD38 _- Cells, display CD34 ⁺ CD38 ^- CD33, CD123, CD45RA, CD44, CD117, HLA-DR and cocktail expression ratios on cells, CD34 was calculated ⁺ CD38 _- CD33 ⁺ Cells, CD34 ⁺ CD38 _- CD123 ⁺ Cells, CD34 ⁺ CD38 ^- CD45RA ⁺ Cells, CD34 ⁺ CD38 ^- CD44 ⁺ Cells, CD34 ⁺ CD38 ^- CD117 ⁺ Cells, CD34 ⁺ CD38 _- HLA-DR ⁺ Cells and CD34 ⁺ CD38 _- cocktail ⁺ The proportion of cells occupying the nucleus;

b2 Using CD45/CD117 spot diagram to distinguish CD117 in R1 according to CD117 expression or not ⁺ Cells, using a CD117/CD34 dot map, distinguish CD117 in R8 based on whether CD34 is expressed or not ⁺ CD34 _- Cells, displaying CD117 ⁺ CD34 ^- The expression ratios of CD33, CD123, CD45RA, CD44, CD117, HLA-DR and cocktail on the cells were calculated to calculate CD117 ⁺ CD34 ^- CD33 ⁺ Cell, CD117 ⁺ CD34 ^- CD123 ⁺ Cell, CD117 ⁺ CD34 ^- CD45RA ⁺ Cell, CD117 ⁺ CD34 ^- CD44 ⁺ Cell, CD117 ⁺ CD34 ^- CD117 ⁺ Cell, CD117 ⁺ CD34 ^- HLA-DR ⁺ Cells and CD117 ⁺ CD34 ^- cocktail ⁺ The proportion of cells occupying the nucleus;

the mixed antibodies are CD7, CD11b, CD22, CD56, CLL-1 and TIM-3.