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

Abstract

The present invention provides a lactic acid antigen and antibody at lysine 388 of human NBS1 protein, and a preparation method and application thereof, which belong to the field of biomedical technology. The present invention provides a lactic acid antigen peptide at lysine 388 of human NBS1 protein, and uses the antigen peptide to prepare an antibody against lactic acid at lysine 388 of human NBS1 protein. The antibody against lactic acid at lysine 388 of human NBS1 protein of the present invention can detect the expression difference between normal cells, tumor cells and tumor cells after medication, which is helpful to study the role of lactic acid modification of human NBS1 protein in the occurrence and development of tumor diseases, and provide potential targets for the diagnosis or treatment of clinical tumor diseases; the antibody against lactic acid at lysine 388 of human NBS1 protein of the present invention can detect the lactic acid 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.

Description

Technical Field

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

[0002] The MRN (MRE11-RAD50-NBS1) complex plays a crucial role in the perception and repair of DNA damage. Double-strand breaks are considered one of the most lethal forms of DNA damage, as one 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 types of cancer. Analysis of surgically resected gastric cancer specimens after chemotherapy showed that low expression levels of the MRN complex were associated with a strong response to chemotherapy and surgical resection. In vitro experiments have shown that disruption of the MRN complex or its components can confer sensitivity to cisplatin on cancer cells, 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 localizes to double-strand break sites as a damage sensor and recruits the key sensor ATM kinase to the double-strand break sites, amplifying and transducing the signal to numerous downstream effectors. In tumorigenesis studies, NBS1 mutations have been observed in breast cancer, prostate cancer, lung cancer, liver cancer, and intrahepatic cholangiocarcinoma and are associated with increased cancer susceptibility. In tumor progression mechanism studies, NBS1 induces the expression of heat shock protein family members HSPA4 or HSPA14, thereby inducing tumor migration, invasion, and transformation activities. In chemoresistance mechanism studies, downregulated expression of NBS1 is associated with hypoxia-induced chemoresistance, inhibition of double-strand break repair pathways, and p53 activation in medulloblastoma cells. In radioresistance mechanism studies, NBS1 can stabilize hypoxia-inducible factors and promote cancer cell migration and invasion under ionizing radiation.

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

[0005] In view of the above problems, the object of the present invention is to provide a human NBS1 protein lysine 388 site lactic acid antigen, antibody and preparation method and application thereof.

[0006] 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 lactic acid.

[0007] The present invention also provides an antibody against the lactic acid at lysine 388 of human NBS1 protein. The antibody is prepared by immunizing an animal with the lactic acid antigen peptide at lysine 388 of human NBS1 protein as an immunogen.

[0008] The present invention also provides a method for preparing the lactic acid antibody against human NBS1 protein lysine 388, comprising the following steps:

[0009] S1. coupling the N-terminus of the antigen peptide with a carrier protein to obtain an antigen;

[0010] S2. Immunize animals with the antigen from S1 and collect antiserum;

[0011] S3. Screening and purifying the antiserum to obtain the lactic acid antibody against lysine 388 site of human NBS1 protein.

[0012] Preferably, the carrier protein comprises KLH.

[0013] Preferably, the immunized animal comprises a rabbit.

[0014] As a preferred embodiment of the preparation method of the present invention, the immunization in step S2 adopts two points of subcutaneous injection of antigens on both shoulders and two points of muscle on both hind legs; the number of immunizations is 4 times, including the 1st, 21st, 35th and 54th days; 4 blood draws are performed after the third immunization, namely on the 45th, 50th, 64th and 69th days after the first immunization.

[0015] 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 ELISA experiment, and purifying the antiserum by affinity separation-affinity purification cycle technology.

[0016] 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 (antigenic 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.

[0017] The present invention also provides the use of the human NBS1 protein lysine 388 site lactic acid antigen peptide in the preparation of a preparation for detecting the human NBS1 protein lysine 388 site lactic acid.

[0018] The present invention also provides use of the antibody against lactic acid at lysine 388 of human NBS1 protein in the preparation of a preparation for detecting lactic acid at lysine 388 of human NBS1 protein.

[0019] The present invention also provides the use of the lactic acid antibody against lysine 388 of human NBS1 protein in the preparation of products for diagnosis, efficacy assessment or prognosis assessment of tumors.

[0020] The present invention also provides a kit for diagnosing, judging the therapeutic effect or determining the prognosis of a tumor, the kit comprising the lactated antigenic peptide at lysine 388 of human NBS1 protein or the lactated antibody against lysine 388 of human NBS1 protein.

[0021] 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 against lactated human NBS1 protein at lysine 388. The antibody against lactated human NBS1 protein at lysine 388 of the present invention can detect the expression difference between normal cells, tumor cells and tumor cells after medication, which is helpful to study the role of lactated modification of human NBS1 protein in the occurrence and development of tumor diseases, and provide potential targets for the diagnosis or treatment of clinical tumor diseases; the antibody against lactated human NBS1 protein at lysine 388 of the present invention can detect the lactated level of human NBS1 protein, explore its relationship with tumor diagnosis and chemoradiotherapy resistance, and has broad clinical application prospects in disease diagnosis, treatment and prognosis determination. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 This is a diagram of the NBS1 protein structure.

[0023] Figure 2This is a technical roadmap for preparing and purifying antibodies against the lactic acid esterification of the human NBS1 protein K388 site according to an embodiment of the present invention.

[0024] Figure 3 The lactylation modification mass spectrometry was used to detect the lactylation of NBS1 protein.

[0025] Figure 4 The figure shows the results of identifying the NBS1-K388la antibody using a dot blot experiment.

[0026] Figure 5 This is a result diagram of the immunohistochemistry experiment to identify the specificity of the antibody.

[0027] Figure 6 The results of prognostic analysis of the antibody against the lactic acid site of human NBS1 protein K388, where 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 lactic acid site of human NBS1 protein K388. DETAILED DESCRIPTION

[0028] In order to more concisely and clearly demonstrate the technical solution, purpose and advantages of the present invention, the technical solution of the present invention is described in detail below in conjunction with specific embodiments and drawings.

[0029] See also Figure 1 , Figure 2 According to the human NBS1 protein structure partition diagram, the present invention provides a method for preparing and applying a lactic acid antibody against the K388 site of the human NBS1 protein, comprising the following steps:

[0030] (1) Preparation of human NBS1 protein K388 site-specific lactylation antibody;

[0031] (2) Purification of antibodies specific for the lactylation site of human NBS1 protein K388;

[0032] (3) Identification and preservation of antibodies specific for lactylation of the K388 site of human NBS1 protein.

[0033] The following is a further description of the technical route for preparing the human NBS1 protein K388 site-specific lactic acid antibody provided by the present invention in combination with specific examples and with reference to the accompanying drawings.

[0034] Example 1 Determination of lactylation sites of human NBS1 protein

[0035] The specific method is as follows:

[0036] (1) We first screened the possible lactylated sites in human NBS1 protein using bioinformatics software NetPhorest 2.0 and NetPhos 2.0, and then confirmed by mass spectrometry that K388 is a lactylated site in the amino acid sequence of human NBS1 ( Figure 3 ).

[0037] (2) Design of lactylated antigenic peptide at K388 of human NBS1 protein. With K388 of human NBS1 protein as the center, four NBS1 amino acid sequences adjacent to the N-terminus and five NBS1 amino acid sequences connected to the C-terminus, the designed synthetic peptide (antigenic peptide) sequence adds a lactylated group to amino acid K388. The peptide sequence is: KEIKVS-(lactyl)K-MEQKFR.

[0038] Example 2 Synthesis of antigenic peptides containing K388 lactylation site and animal immunization The specific methods are as follows:

[0039] (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), and 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 an affinity purification chromatography column (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 an affinity separation chromatography column (K388-BSA). The synthetic peptides were all tested by Mass Spectral, and the results are as follows: Figure 5 shown.

[0040] (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, and the blood was compressed with a cotton ball to stop bleeding. The blood was placed at room temperature for about 1 hour to allow the blood to coagulate and form a blood clot. The blood was placed at 4°C for 2 hours to allow the serum to precipitate. The blood was centrifuged at 2500 g for 10 minutes, and the supernatant was aspirated and marked as negative control serum. The blood was aliquoted and stored at -20°C for testing.

[0041] (3) Animal immunization:

[0042] a. Dissolve K388la-KLH synthetic peptide powders in sterile 1×PBS, draw antigen solution with one sterile syringe, draw an equal amount of Freund's complete adjuvant (CFA) with another syringe, connect the two with a plastic tube, and pump back and forth repeatedly to mix the K388la-KLH peptide segments with CFA until a completely emulsified emulsion is formed, which does not spread when dropped into water;

[0043] b. For the first immunization, the antigen emulsion is injected at two points under the skin of both shoulders and two points in the muscles of both hind legs. The total amount of the two antigens for the first immunization is about 0.61mg;

[0044] c. The first, second and third booster immunizations were carried out 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 injected in the same manner as the first immunization. The total amount of antigen in the booster immunization was about 0.9 mg.

[0045] d. Take blood samples 45, 50, 64, and 69 days after the first immunization and collect a large amount of serum. Seal the beaker containing the collected blood and let it stand overnight at room temperature to shrink the blood clot. The next day, aseptically dispense the precipitated serum into 50 mL centrifuge tubes, centrifuge at 4000 g for 10 min, take the supernatant, dispense 1 mL / tube, mark as post-immunization antiserum, and store at -20°C.

[0046] Example 3 Screening and purification of human NBS1 protein K388 site-specific lactylation antibodies

[0047] (1) Determination and screening of the titer of antiserum after immunization, the specific steps are as follows:

[0048] a. Coat the lactated antigen K388la-BSA and non-lactated antigen K388-BSA with antigen coating solution (CBS), add 0.1 mL to each well of a 96-well ELISA plate, cover the plate, shake and mix, and coat overnight at 4°C (more than 12 hours);

[0049] b. After coating, discard the liquid in the wells, wash each well of the plate with 1×PBST, discard the washing solution, repeat 3 times, and wipe off the residual liquid on the filter paper after each wash. Add blocking buffer (0.25% BSA / PBST) to each well, 200μL / well, incubate at 37℃ for 2h, wash the plate 3 times with 1×PBST, and wipe off the residual liquid on the filter paper after each wash;

[0050] c. Dilute the post-immune antiserum with 1×PBST at a ratio of 1:1000, 1:3000, 1:9000, 1:27000, 1:81000, 1:243000, and 1:729000, respectively, with 1×PBS buffer as a blank control, and add them to a 96-well ELISA plate, cover the plate, and incubate at 37°C for 1 hour;

[0051] d. Discard the liquid in the wells and wash the plate 3 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℃ for 1h. Cover the plate and incubate at 37℃ for 1h. Wash the plate 5 times with 1×PBST and dry. Add temporarily prepared TMB colorimetric solution, 100μL / well, and react at room temperature in the dark for 30min. Add 2MH2SO4 to terminate the reaction, 50μL / well. Measure the OD value of each well with an enzyme reader at 450nm;

[0052] e. The ELISA test values ​​of the antiserum 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 subsequently purified. The screening results are shown in Table 1 below.

[0053] Table 1 OD values ​​of antiserum ELISA after immunization with K388 antigen

[0054] Original serum dilution K388-BSA K388la-BSA 1:1000 2.689 2.698 1:3000 2.235 2.242 1:9000 1.523 1.862 1:27000 0.529 0.758 1:81000 0.136 0.192 1:243000 0.025 0.036 1:729000 0.012 0.015 BLANK 0.002 0.009

[0055] (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:

[0056] a. Dissolve K388-BSA and K388la-BSA in coupling buffer to a concentration of 0.7 mg / mL. Take 7 mg of K388-BSA and 6 mg of K388la-BSA solution and add them to 5 mL of resin, mix well and add them to the chromatography column and mix well at room temperature for 15 minutes. Place the chromatography column upright at room temperature for 30 minutes, remove the caps at the upper and lower ends of the chromatography column, collect the effluent, and rinse the column with 3 times the volume of the resin coupling buffer;

[0057] b. Cover the bottom cover 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 stand for 30 minutes.

[0058] 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 cover, add 1 volume of storage buffer, and store at 4 degrees for later use.

[0059] (3) Purification of lactic acid antibody: Antibody purification was prepared using a Thermo Scientific Coupling Resin kit. The specific steps are as follows:

[0060] a. Remove the cap at the bottom of the chromatography column, release the storage buffer, and add 6 mL of binding buffer to wash the chromatography column. Add K388 antiserum to the corresponding non-lactated chromatography column, collect the effluent, repeat twice to obtain K388 effluent, then wash the chromatography column with 12 mL of effluent, retain the eluate at the bottom, and finally elute the chromatography column with the eluate to obtain the corresponding non-lactated antibody;

[0061] b. Add the effluent collected in a to the corresponding washed lactic acid column, then wash the column with 12 mL of washing solution, and finally elute the column with eluent to obtain the corresponding K388 lactic acid antibody. The antibody is dissolved in 1×PBS and 0.1% NaN3 is added for later use.

[0062] Example 4 Application of the human NBS1 protein K388 site lactic acid antibody to test the sensitivity of radiotherapy and chemotherapy

[0063] (1) Dot blot kit (Cat. No.: IEMed-K236) was used to detect the lactic acid peptide specificity of the purified antibody: synthetic non-lactated NBS1 peptide (control peptide) and NBS1 peptide with lactic acid modification at amino acid position 388 (modified peptide) were used as antigens (modified peptide and control peptide) and dropped on cellulose acetate membrane at 1, 4, 16, and 64 ng / drop, respectively, and air-dried. Then, polyclonal antibodies with gradient dilutions were used to react with them for dot blot experiments.

[0064] The results are as follows Figure 4 As shown, the polyclonal antibody against NBS1 with lactylation at amino acid position 388 can recognize the polypeptide sequence against NBS1 with lactylation at amino acid position 388, but does not recognize the NBS1 polypeptide sequence without lactylation, and the antibody titer is >1:100000.

[0065] (2) Immunohistochemistry to identify antibody specificity

[0066] 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 lactic acid-modified peptide NBS1 peptide segment, and the other section was not treated with the lactic acid-modified peptide NBS1 peptide segment. Then, immunohistochemistry experiments were performed. The specific experimental steps are as follows:

[0067] Tissue fixation: Before dewaxing, bake the sections in a thermostat at 60°C for 60 min.

[0068] Dewaxing: Soak the sections in xylene for 15 minutes, then replace with new xylene and soak for another 15 minutes.

[0069] 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.

[0070] Washing: Soak in dd H2O for 5 min, wash 3 times.

[0071] Antigen repair (microwave radiation method): 0.3% H2O2 methanol treatment for 10 minutes, rinse with tap water and then with distilled water, put the slices into a special repair box with pH 6.0 citrate buffer and heat to boiling for 10 minutes. After the repair solution cools to room temperature, wash with PBS 3 times.

[0072] Washing: soak in dd H2O for 5 min, wash twice, soak in PBST for 5 min, wash twice.

[0073] Inactivate enzyme: Soak the slices in 20 ml of inactivate enzyme reagent, protect from light, and leave at room temperature for 15 minutes.

[0074] Washing: Soak in PBST for 5 min, wash 3 times.

[0075] Blocking: Add appropriate amount of blocking solution and incubate in a wet box at room temperature for 60 minutes.

[0076] 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.

[0077] Rewarming: After removing from 4°C, incubate at room temperature for 60 min.

[0078] Washing: Rinse gently with PBST and soak for 5 minutes, wash 4 times.

[0079] Enzyme-labeled secondary antibody reaction: Add diluted HRP-labeled secondary antibody to each tissue and incubate at room temperature for 60 minutes. Washing: Gently rinse with PBST and soak for 5 minutes, wash 4 times.

[0080] Colorimetric DAB method: prepare DAB colorimetric solution, react in the dark for 10-15 minutes, add it dropwise onto the slices, and color for 1-5 minutes.

[0081] Stop color development: Use distilled water to stop the color development reaction.

[0082] Restaining: Add an appropriate amount of hematoxylin solution to each tissue, stain for 5-10 minutes, and rinse with distilled water.

[0083] 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.

[0084] Sealing: 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 with xylene and soak for another 15 minutes, then cover with a glass slide.

[0085] The results are as follows Figure 5 As shown, the specificity of the NBS1 K3881a antibody is good and can be used for immunohistochemistry.

[0086] Example 5 The specific method for prognostic analysis of human NBS1 protein K388 site lactic acid antibody is as follows:

[0087] (1) The tumor sections of gastric cancer patients were stained using the immunohistochemical method in Example 4, and the sections were scored. The scoring method is as follows: the score of the staining intensity is defined as: negative 0 point; weak 1 point; moderate 2 points; strong positive 3 points. The frequency of positive cells is defined as: less than 5%, 0 point: 5%-25%, 1 point: 26%-50%, 2 points: 51%-75%, 3 points: greater than 75%, 4 points. The score of the staining intensity is multiplied by the frequency score of the positive cells to obtain the final immunohistochemical score.

[0088] (2) According to the immunohistochemical score, the tumor tissues were divided into high and low expression groups, and the prognostic analysis was performed based on the patients' prognostic information: 94 gastric cancer patients' tumor tissues were collected, immunohistochemical staining was performed using NBS1 K388la antibody, and the patients' prognostic analysis was performed. The results are as follows: Figure 6 As shown, the higher the expression level of NBS1 protein lactylation modification, the worse the prognosis of tumor patients.

[0089] Finally, it should be noted that the above embodiments are only used to illustrate the technical solution of the present invention rather than to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that the technical solution of the present invention can be modified or replaced by equivalents without departing from the essence and scope of the technical solution of the present invention.

Claims

1. A lactic acid 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 against the lactated antigen peptide at lysine 388 of human NBS1 protein, characterized in that: The antibody is prepared by immunizing an animal with the lactated antigen peptide at lysine 388 of human NBS1 protein as claimed in claim 1 as an immunogen, and purifying the antiserum using affinity separation-affinity purification cycle technology; the immunized animal is a rabbit.

3. The method for preparing the antibody against the lactated antigen peptide at lysine 388 of human NBS1 protein according to claim 2, characterized in that: The following steps are involved: S1. coupling the N-terminus of the antigen peptide according to claim 1 with a carrier protein, Get antigen; S2. Immunize animals with the antigen from S1 and collect antiserum; S3. Screening and purifying the antiserum to obtain the antibody against the lactated antigen peptide at lysine 388 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 under the skin of both shoulders and two points in the muscles of 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.

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

6. Use of the lactated antigen peptide at lysine 388 of human NBS1 protein according to claim 1 in preparing polyclonal antibodies.

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

8. Use of the antibody against the lactated antigen peptide at lysine 388 of human NBS1 protein according to claim 2 in the preparation of a product for diagnosis, efficacy assessment or prognosis assessment of tumors, characterized in that: The tumor is gastric cancer.

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