Human NBS1 protein antigen lactylated at lysine 388, antibody, preparation method therefor, and use thereof

Abstract

Provided are a human NBS1 protein antigen lactylated at lysine 388, an antibody, a preparation method therefor, and a use thereof, relating to the technical field of biomedicine. A human NBS1 protein antigen peptide lactylated at lysine 388 is provided, and is used to prepare an antibody targeting lactylation at lysine 388 of human NBS1 protein. The antibody targeting lactylation at lysine 388 of human NBS1 protein can detect expression differences among normal cells, tumor cells, and drug-treated tumor cells, thereby helping to study the role of lactylation modification of human NBS1 protein in the occurrence and development of tumor diseases, and providing potential therapeutic targets for clinical diagnosis or treatment of tumor diseases. The antibody targeting lactylation at lysine 388 of human NBS1 protein can be used to detect the lactylation level of human NBS1 protein, and to investigate its relationship with tumor diagnosis and resistance to radiotherapy and chemotherapy, thereby offering broad clinical application prospects in disease diagnosis, treatment, and prognosis evaluation.

Description

A human NBS1 protein lysine 388 site lactic acid antigen, antibody and preparation method and application thereof Technical Field

[0001] The present invention belongs to the field of biomedicine technology, and specifically relates to a human NBS1 protein lysine 388 site lactated antigen, antibody, and preparation method and application thereof. Background Art

[0002] The MRN (MRE11-RAD50-NBS1) complex plays a crucial role in the sensing and repair of DNA damage. Double-strand breaks are considered one of the most lethal forms of DNA damage, as a single unrepaired double-strand break is sufficient to cause permanent growth arrest or cell death. The MRN complex maintains genomic stability and prevents malignant transformation of normal cells. Its function in cancer development and its potential as an anticancer target have been extensively explored in various cancer types. Analysis of surgically resected gastric cancer specimens after chemotherapy revealed that low expression levels of the MRN complex are associated with a robust response to chemotherapy and surgical resection. In vitro experiments have shown that disruption of the MRN complex or its components can sensitize cancer cells to cisplatin, along with increased DNA damage and cytotoxicity. Overexpression of the MRN complex can confer radioresistance to rectal cancer.

[0003] NBS1 is a multifunctional protein that acts as a damage sensor localized to double-strand break sites, recruiting the key sensor ATM kinase to the sites of double-strand breaks, amplifying and transducing the signal to numerous downstream effectors. In tumorigenesis research, NBS1 mutations have been observed in breast cancer, prostate cancer, lung cancer, liver cancer, and intrahepatic bile duct carcinoma, and are associated with increased cancer susceptibility. In studies of tumor progression mechanisms, NBS1 induces the expression of heat shock protein family members HSPA4 or HSPA14, thereby inducing tumor migration, invasion, and transformation activities. In studies of chemoresistance mechanisms, downregulated NBS1 expression is associated with hypoxia-induced chemoresistance, inhibition of the double-strand break repair pathway, and p53 activation in medulloblastoma cells. In studies of radioresistance mechanisms, NBS1 can stabilize hypoxia-inducible factors and promote cancer cell migration and invasion in response to ionizing radiation.

[0004] Post-translational modifications such as lactylation, phosphorylation, and ubiquitination play key roles in physiological and pathological processes mediated by cell signaling pathways. Given the crucial role of NBS1 in tumorigenesis and progression, further investigation of post-translational modifications of the NBS1 protein is of great significance.

[0005] Summary of the Invention

[0006] In view of the above problems, the purpose of the present invention is to provide a human NBS1 protein lysine 388 site lactation antigen, antibody and preparation method and application thereof.

[0007] To achieve the above object, the technical solution adopted by the present invention is: a lactated antigenic peptide at lysine 388 of human NBS1 protein, the active amino acid sequence of the antigenic peptide is shown in SEQ ID NO: 1, and the lysine therein is modified by lactation.

[0008] The present invention also provides an antibody against the lactation of human NBS1 protein at lysine 388, wherein the antibody is prepared by immunizing an animal with the lactation antigen peptide at lysine 388 of human NBS1 protein as an immunogen.

[0009] The present invention also provides a method for preparing the antibody against the lactation of human NBS1 protein at lysine 388, comprising the following steps:

[0010] S1. coupling the N-terminus of the antigen peptide with a carrier protein to obtain an antigen;

[0011] S2. Immunize animals with the antigen from S1 and collect antiserum;

[0012] S3. Screening and purifying the antiserum to obtain the lactic acid antibody against lysine 388 site of human NBS1 protein.

[0013] Preferably, the carrier protein comprises hemocyanin (KLH).

[0014] Preferably, the immunized animal includes a rabbit.

[0015] As a preferred embodiment of the preparation method of the present invention, the immunization in step S2 adopts the injection of antigens at two points of the subcutaneous tissue on both shoulders and two points of the muscles on both hind legs; the number of immunizations is 4 times, including the 1st, 21st, 35th and 54th days; and blood is collected 4 times after the third immunization, namely on the 45th, 50th, 64th and 69th days after the first immunization.

[0016] As a preferred embodiment of the preparation method of the present invention, the screening and purification in step S3 specifically comprises determining the titer of the collected antiserum against the lactated antigen peptide at lysine 388 of the human NBS1 protein according to claim 1 by an ELISA experiment, and purifying the antiserum by affinity separation-affinity purification cycle technology.

[0017] Preferably, the affinity separation-affinity purification cycle technology is used to purify the antiserum as follows: affinity chromatography using a synthetic peptide coupled to a carrier protein bovine serum albumin (BSA) and an agarose chromatography column is used for antibody affinity separation and affinity purification; non-lactated synthetic peptides are used to couple bovine serum albumin (BSA) and agarose as fillers for the chromatography column to perform affinity separation to remove non-lactated antibodies in the antiserum, thereby obtaining an effluent containing lactated antibodies; and then lactated synthetic peptides (antigen peptides) are used to couple bovine serum albumin (BSA) and agarose as fillers for the chromatography column to perform affinity purification to remove low-affinity, low-sequence-complexity epitope antibodies in the lactated antibodies.

[0018] The present invention also provides use of the human NBS1 protein lysine 388 lactation antigen peptide in preparing a preparation for detecting human NBS1 protein lysine 388 lactation.

[0019] The present invention also provides use of the antibody against lactation at lysine 388 of human NBS1 protein in preparing a preparation for detecting lactation at lysine 388 of human NBS1 protein.

[0020] The present invention also provides the use of the antibody targeting the lactated site of lysine 388 of human NBS1 protein in the preparation of a product for the diagnosis, efficacy assessment or prognosis determination of tumors.

[0021] The present invention also provides a kit for diagnosing, judging the therapeutic effect or determining the prognosis of a tumor, comprising the human NBS1 protein lysine 388 lactated antigen peptide or the antibody against human NBS1 protein lysine 388 lactated antibody.

[0022] The beneficial effects of the present invention are as follows: the present invention provides a lactated antigenic peptide at lysine 388 of human NBS1 protein, and uses the antigenic peptide to prepare an antibody targeting lactated antibody at lysine 388 of human NBS1 protein. The antibody targeting lactated antibody at lysine 388 of human NBS1 protein of the present invention can detect the expression differences between normal cells, tumor cells and tumor cells after drug administration, which is helpful for studying the role of lactated modification of human NBS1 protein in the occurrence and development of tumor diseases, and provides a potential target for the diagnosis or treatment of clinical tumor diseases; the antibody targeting lactated antibody at lysine 388 of human NBS1 protein of the present invention can detect the lactated level of human NBS1 protein, explore its relationship with tumor diagnosis and radiotherapy and chemotherapy resistance, and has broad clinical application prospects in disease diagnosis, treatment and prognosis determination. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 is a diagram of the NBS1 protein structure.

[0024] FIG2 is a technical roadmap for preparing and purifying antibodies against the lactated K388 site of human NBS1 protein according to an embodiment of the present invention.

[0025] Figure 3 shows the detection of NBS1 protein lactylation by lactylation modification mass spectrometry.

[0026] FIG4 is a graph showing the results of identifying the NBS1-K3881a antibody using a dot blot assay.

[0027] FIG5 is a graph showing the results of immunohistochemistry experiments to identify antibody specificity.

[0028] Figure 6 shows the prognostic analysis results of the antibody against the lactate at K388 of the human NBS1 protein, wherein A is a typical immunohistochemical image of tumor tissues collected from 94 gastric cancer patients for immunohistochemical staining; B is the survival prognosis result of the antibody against the lactate at K388 of the human NBS1 protein. DETAILED DESCRIPTION

[0029] In order to more concisely and clearly demonstrate the technical solutions, objectives and advantages of the present invention, the technical solutions of the present invention are described in detail below with reference to specific embodiments and accompanying drawings.

[0030] 1 and 2 , the present invention provides a method for preparing and applying an antibody targeting the lactic acid residue at the K388 site of the human NBS1 protein according to the structural partition diagram of the human NBS1 protein, comprising the following steps:

[0031] (1) Preparation of antibodies specific for the lactylation site of human NBS1 protein K388;

[0032] (2) Purification of antibodies specific for the lactylation site at K388 of the human NBS1 protein;

[0033] (3) Identification and preservation of antibodies specific for the lactylation site K388 of human NBS1 protein.

[0034] The following is a further description of the technical route for preparing the human NBS1 protein K388 site-specific lactylation antibody provided by the present invention in combination with specific examples and with reference to the accompanying drawings.

[0035] Example 1 Determination of the lactylation site of human NBS1 protein

[0036] The specific method is as follows:

[0037] (1) We first screened potential lactylation sites in human NBS1 protein using the bioinformatics software NetPhorest2.0 and NetPhos2.0, and then confirmed by mass spectrometry that K388 was a lactylation site in the amino acid sequence of human NBS1 (Figure 3).

[0038] (2) Design of a lactated antigenic peptide targeting the K388 site of human NBS1 protein. Centered on the K388 site of human NBS1 protein, with the N-terminus adjacent to four NBS1 amino acid sequences and the C-terminus connected to five NBS1 amino acid sequences, a synthetic peptide (antigenic peptide) sequence was designed, with a lactyl group added to amino acid K388. The peptide sequence was: KEIKVS-(lactyl)K-MEQKFR.

[0039] Example 2 Synthesis of antigenic peptides containing K388 lactylation site and animal immunization

[0040] The specific method is as follows:

[0041] (1) According to the design of the hapten synthetic peptide, a lactated group was added to the K388 site to obtain a lactated synthetic peptide (antigenic peptide), which was then coupled with hemocyanin (KLH) to obtain a complete antigen for rabbit immunization (K388la-KLH). The lactated synthetic peptide (antigenic peptide) was coupled with bovine serum albumin (BSA) by the glutaraldehyde method and used as a filler for affinity purification chromatography columns (K388la-BSA). Correspondingly, a non-lactated synthetic peptide corresponding to K388 was synthesized and coupled with bovine serum albumin (BSA) to be used as a filler for affinity separation chromatography columns (K388-BSA). The synthetic peptides were all detected by Mass Spectral, and the results are shown in Figure 5.

[0042] (2) Preparation of negative serum: 3 mL of blood was collected from the ear vein of a New Zealand white rabbit (2-3 kg, female, strong, purchased from the Slack Laboratory Animal Center of the Chinese Academy of Sciences) for injection into a blood collection tube. Hemostasis was achieved by pressing with a cotton ball. The blood was allowed to stand at room temperature for about 1 hour to form a clot. The serum was then allowed to precipitate at 4°C for 2 hours. The serum was centrifuged at 2500 g for 10 minutes. The supernatant was aspirated and labeled as negative control serum. The serum was aliquoted and stored at -20°C until testing.

[0043] (3) Animal immunization:

[0044] Dissolve the K388la-KLH synthetic peptide powder in sterile 1× PBS. Use one sterile syringe to draw up the antigen solution and another syringe to draw up an equal amount of complete Freund's adjuvant (CFA). Connect the two syringes with plastic tubing and repeatedly pump back and forth to mix the K388la-KLH peptide and CFA until a completely emulsified emulsion is formed that does not spread when dropped into water.

[0045] b. For the first immunization, the antigen emulsion is injected at multiple locations: two subcutaneous points on both shoulders and two points on the muscles of both hind legs. The total amount of the two antigens for the first immunization is approximately 0.61 mg.

[0046] c. The first, second, and third booster immunizations were performed 21, 35, and 54 days after the first immunization. Incomplete Freund's adjuvant (IFA) was used instead of CFA as the immune adjuvant to prepare the antigen emulsion and the emulsion was injected as the first immunization. The total amount of antigen in each booster immunization was approximately 0.9 mg.

[0047] d. Collect blood 45, 50, 64, and 69 days after the initial immunization and collect large amounts of serum. Seal the collected blood beaker and let it sit at room temperature overnight to allow the blood clot to shrink. The next day, aseptically aliquot the separated serum into 50 mL centrifuge tubes and centrifuge at 4000 g for 10 minutes. Remove the supernatant and aliquot 1 mL / tube. Label this as post-immunization antiserum and store at -20°C.

[0048] Example 3 Screening and purification of human NBS1 protein K388 site-specific lactylation antibodies

[0049] (1) Determination and screening of the titer of antiserum after immunization. The specific steps are as follows:

[0050] a. Coat the lactated antigen K388la-BSA and the non-lactated antigen K388-BSA with antigen coating solution (CBS) and add 0.1 mL per well to a 96-well ELISA plate. Cover the plate and shake to mix thoroughly. Coat overnight at 4°C (for at least 12 hours).

[0051] b. After coating, discard the liquid in the wells and thoroughly wash each well of the plate with 1× PBST. Discard the wash solution and repeat three times, tapping off any remaining liquid on filter paper after each wash. Add 200 μL / well of blocking buffer (0.25% BSA / PBST) to each well and incubate at 37°C for 2 hours. Wash the plate three times with 1× PBST, tapping off any remaining liquid on filter paper after each wash.

[0052] c. Dilute the post-immune antiserum with 1× PBST at the ratio of 1:1000, 1:3000, 1:9000, 1:27000, 1:81000, 1:243000, and 1:729000, respectively, using 1× PBS buffer as a blank control. Add the diluted serum to a 96-well microtiter plate, seal the plate, and incubate at 37°C for 1 hour.

[0053] d. Discard the liquid in the wells and wash the plate three times with 1×PBST. Add HRP-labeled goat anti-rabbit IgG secondary antibody diluted 1:5000 with 1×PBST, 100μL / well, and incubate at 37°C for 1 hour. Cover the plate and incubate at 37°C for 1 hour. Wash the plate five times with 1×PBST and dry. Add the temporarily prepared TMB color development solution, 100μL / well, and react at room temperature in the dark for 30 minutes. Add 2M H2SO4 to stop the reaction, 50μL / well. Measure the OD value of each well at 450nm using a microplate reader;

[0054] e. The ELISA test values ​​of the antisera after immunization with K388 antigen at dilutions of 1:1000, 1:3000, and 1:9000 were all above 1.0. The screening serum (OD450>1.0) reaching 1:10,000 was considered to meet the standard and could be used for subsequent purification. The screening results are shown in Table 1 below.

[0055] Table 1 OD values ​​of antiserum ELISA after immunization with K388 antigen

[0056] (2) Preparation of lactated synthetic peptide chromatography column and non-lactated synthetic peptide chromatography column. The synthetic peptide coupling chromatography column was prepared using a Thermo Scientific Coupling Resin kit. The specific steps are as follows:

[0057] a. Dissolve K388-BSA and K388la-BSA in coupling buffer to a concentration of 0.7 mg / mL each. Add 7 mg of K388-BSA and 6 mg of K388la-BSA to 5 mL of resin, mix thoroughly, and then add each to the chromatography column. Mix thoroughly at room temperature for 15 minutes. Leave the column upright at room temperature for 30 minutes. Remove the caps at the top and bottom of the column, collect the runoff, and rinse the column with 3 volumes of coupling buffer equal to the volume of resin.

[0058] b. Cover the bottom of the chromatography column, add 50mM L-Cysteine·HCl to the coupling buffer, mix well, and then add an equal volume of buffer to the chromatography column. Mix well at room temperature for 15 minutes and let it stand for 30 minutes.

[0059] c. Remove the bottom cover to release the coupling buffer, wash the column with 6 volumes of washing solution (1M NaCl), then rinse the column with 2 volumes of storage buffer, cover the column, add 1 volume of storage buffer, and store at 4 degrees for later use.

[0060] (3) Purification of lactated antibodies: Antibody purification was performed using a Thermo Scientific Coupling Resin kit. The specific steps are as follows:

[0061] a. Remove the cap from the bottom of the chromatography column to release the storage buffer. Add 6 mL of binding buffer to wash the column. Add the K388 antiserum to the corresponding non-lactated chromatography column and collect the effluent. Repeat twice to obtain the K388 effluent. Then, wash the chromatography column with 12 mL of rinsing buffer, retaining the eluate at the bottom. Finally, elute the chromatography column with the eluate to obtain the corresponding non-lactated antibody.

[0062] b. Add the effluent collected in step a to the corresponding rinsed lactated column, then wash the column with 12 mL of washing solution, and finally elute the column with eluent to obtain the corresponding K388 lactated antibody. The antibody was dissolved in 1× PBS and 0.1% NaN3 was added for later use.

[0063] Example 4: Application of the human NBS1 protein K388 lactate antibody to test the sensitivity of radiotherapy and chemotherapy

[0064] (1) Dot blot kit (Cat. No.: IEMed-K236) was used to detect the lactic acid peptide specificity of the purified antibody: a synthetic non-lactated NBS1 peptide (control peptide) and a peptide with lactic acid modification at amino acid position 388 of NBS1 (modified peptide) were used as antigens and dropped onto cellulose acetate membranes at 1, 4, 16, and 64 ng / drop, respectively, and air-dried. Dot blot experiments were then performed using polyclonal antibodies diluted in a gradient concentration to react with them.

[0065] The results are shown in Figure 4 . The polyclonal antibody against NBS1 with lactylation at amino acid position 388 can recognize the polypeptide sequence of NBS1 with lactylation at amino acid position 388, but does not recognize the NBS1 polypeptide sequence without lactylation, and the antibody titer is >1:100,000.

[0066] (2) Immunohistochemistry to identify antibody specificity

[0067] The specificity of the antibody was identified by immunohistochemistry experiments. To determine whether the antibody could be used in immunohistochemistry experiments, two immunohistochemistry sections of gastric cancer tumor tissue were collected. One section was treated with the lactated peptide NBS1 peptide, and the other section was not treated with the lactated peptide NBS1 peptide. Immunohistochemistry experiments were then performed. The specific experimental steps are as follows:

[0068] Tissue fixation: Before dewaxing, bake the sections in a thermostat at 60°C for 60 min.

[0069] Dewaxing: Soak the sections in xylene for 15 minutes, then replace with new xylene and soak for another 15 minutes.

[0070] Hydration: Soak in anhydrous ethanol for 5 minutes, replace with new anhydrous ethanol and soak for another 5 minutes, soak in 95% ethanol for 5 minutes, soak in 85% ethanol for 5 minutes, and soak in 75% ethanol for 5 minutes.

[0071] Washing: Soak in dd H2O for 5 minutes, wash 3 times.

[0072] Antigen retrieval (microwave irradiation): Treat with 0.3% H₂O₂ in methanol for 10 minutes. Rinse with tap water and then distilled water. Place the sections in a dedicated retrieval chamber containing pH 6.0 citrate buffer and heat to boiling for 10 minutes. After the retrieval solution cools to room temperature, wash three times with PBS.

[0073] Washing: soak in dd H2O for 5 minutes, wash twice, soak in PBST for 5 minutes, wash twice.

[0074] Inactivate enzyme: Soak the slices in 20 ml of inactivate enzyme reagent, protect from light, and allow to act at room temperature for 15 minutes.

[0075] Washing: Soak in PBST for 5 minutes, wash three times.

[0076] Blocking: Add appropriate amount of blocking solution and incubate in a wet box at room temperature for 60 minutes.

[0077] Primary antibody reaction: Use neutral gum to draw circles to form a groove, add diluted primary antibody NBS1K388la to each tissue, and incubate overnight in a 4°C wet box.

[0078] Rewarming: After removing from 4°C, incubate at room temperature for 60 minutes.

[0079] Washing: Rinse gently with PBST and soak for 5 minutes, wash 4 times.

[0080] Enzyme-labeled secondary antibody reaction: Add diluted HRP-labeled secondary antibody to each tissue and incubate at room temperature for 60 minutes.

[0081] Washing: Rinse gently with PBST and soak for 5 minutes, wash 4 times.

[0082] Colorimetric DAB method: Prepare DAB colorimetric solution, react in the dark for 10-15 minutes, add it dropwise to the slice, and color for 1-5 minutes.

[0083] Stop color development: stop the color development reaction with distilled water.

[0084] Restaining: Add an appropriate amount of hematoxylin solution to each tissue, stain for 5-10 minutes, and rinse with distilled water.

[0085] Decolorization and anti-blueing: Place the slices in 1% hydrochloric acid-ethanol for 2-3 seconds to decolorize, then quickly take them out and place them in distilled water to stop the decolorization. Then place them in PBST for anti-blueing for 5-10 minutes.

[0086] Seal the slides: Soak in 75% ethanol for 5 minutes, 85% ethanol for 5 minutes, 95% ethanol for 5 minutes, and absolute ethanol for 5 minutes. Soak in xylene for 15 minutes, then replace the xylene and soak for another 15 minutes. Seal the slides with coverslips.

[0087] The results are shown in FIG5 , indicating that the NBS1K3881a antibody has good specificity and can be used for immunohistochemistry.

[0088] Example 5 Prognostic Analysis of Antibodies Targeting K388 Lactation of Human NBS1 Protein

[0089] The specific method is as follows:

[0090] (1) Using the immunohistochemical method described in Example 4, sections of tumors from gastric cancer patients were stained and scored. The scoring method was as follows: the staining intensity score was defined as: negative (0 point); weak (1 point); moderate (2 points); and strongly positive (3 points). The frequency of positive cells was defined as: less than 5%, 0 point: 5%-25%, 1 point: 26%-50%, 2 points: 51%-75%, 3 points: greater than 75%, and 4 points. The staining intensity score was multiplied by the frequency score of positive cells to obtain the final immunohistochemical score.

[0091] (2) The patients were divided into two groups according to immunohistochemical scores, high expression and low expression, and prognostic analysis was performed in combination with the patients' prognostic information: Tumor tissues from 94 gastric cancer patients were collected, immunohistochemically stained with NBS1K388la antibody, and the patients' prognostic analysis was performed. The results are shown in Figure 6. The higher the expression level of NBS1 protein lactylation modification, the worse the prognosis of the tumor patient.

[0092] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that the technical solutions of the present invention may be modified or replaced by equivalents without departing from the essence and scope of the technical solutions of the present invention.

Claims

1. A lactated antigen peptide at lysine 388 of human NBS1 protein, characterized in that: The active amino acid sequence of the antigen peptide is shown in SEQ ID NO: 1, and the lysine therein is modified by lactic acid.

2. An antibody targeting the lactic acid residue at lysine 388 of human NBS1 protein, characterized in that: The antibody is prepared by immunizing an animal with the human NBS1 protein lysine 388 lactated antigen peptide as claimed in claim 1 as an immunogen.

3. The method for preparing the antibody against the lactic acid lysine 388 site of human NBS1 protein according to claim 2, characterized in that: The following steps are involved: S1. The N-terminus of the antigen peptide according to claim 1 is coupled to a carrier protein to obtain an antigen; S2. Immunize animals with the antigen from S1 and collect antiserum; S3. Screening and purifying the antiserum to obtain the lactic acid antibody against lysine 388 site of human NBS1 protein.

4. The preparation method according to claim 3, characterized in that The immunization in step S2 is performed by injecting antigens at two points on the subcutaneous tissue of both shoulders and two points on the muscles of both hind legs; the number of immunizations is 4 times, including on the 1st, 21st, 35th and 54th days; and blood is collected 4 times after the third immunization, namely on the 45th, 50th, 64th and 69th days after the first immunization.

5. The preparation method according to claim 3, characterized in that The screening and purification in step S3 specifically involves determining the titer of the collected antiserum against the lactated antigen peptide at lysine 388 of the human NBS1 protein according to claim 1 by ELISA experiment, and purifying the antiserum by affinity separation-affinity purification cycle technology.

6. Use of the human NBS1 protein lysine 388 lactation antigen peptide according to claim 1 in the preparation of a preparation for detecting human NBS1 protein lysine 388 lactation.

7. Use of the antibody against human NBS1 protein lysine 388 lactation according to claim 2 in the preparation of a preparation for detecting human NBS1 protein lysine 388 lactation.

8. Use of the antibody against human NBS1 protein lysine 388 lactation according to claim 2 in the preparation of a product for tumor diagnosis, efficacy assessment or prognosis determination.

9. A kit for diagnosing, judging therapeutic efficacy or determining prognosis of a tumor, characterized in that: The kit comprises the human NBS1 protein lysine 388 lactation antigen peptide according to claim 1 or the human NBS1 protein lysine 388 lactation antibody according to claim 2.

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