Anti-human ST2 monoclonal antibody and application thereof

Abstract

The invention relates to the technical field of biology, in particular to an anti-human ST2 monoclonal antibody and related application thereof. Pairing antibodies with high affinity and strong specificity are obtained through a mouse hybridoma cell technology, are respectively used for detection and capture, and can be used for detection of clinical samples. The invention provides a method for clinical sample detection, namely time-resolved microsphere immunochromatography detection, the method can be used for quantitative detection of ST2 protein in a clinical sample, and the correlation coefficient is gt; and the test range is 1-250ng / ml, and compared with a commercialized paired antibody, the detection lower limit is superior to that of the commercialized antibody, and the sensitivity is higher. When the paired antibody is used for detecting a clinical sample for 10 times, the CV is less than 15%, and the stability is good.

Description

Technical Field

[0001] The present invention relates to the field of biotechnology, and in particular to a monoclonal antibody against human ST2 and related applications thereof. Background Art

[0002] Interleukin-33 (IL-33) is a member of the IL-1 cytokine family and has dual functions. On the one hand, it can induce the activation of NFkB and MAPK, and on the other hand, it can participate in the regulation of gene expression as a transcription factor. ST2 is a member of the IL-1R superfamily and is mainly expressed in cardiomyocytes and also expressed on a variety of immune cells. The precursor mRNA of ST2 can be divided into three different subtypes after selective splicing: transmembrane ST2L, secretory ST2 (soluble ST2, sST2) and multivariate ST2V (ST2 variant form). ST2L is expressed on immune cells such as Th2 lymphocytes, NK, NKT and mast cells, as well as myeloid cells such as monocytes, dendritic cells and granulocytes. The IL-33 / ST2 cell pathway can promote the immune response of Th1 and Th2, and has a variety of cardiovascular protective effects such as anti-atherosclerosis, anti-myocardial hypertrophy, and anti-myocardial fibrosis. Under mechanical stress, myocardial cells produce a large amount of soluble ST2, which acts as a "decoy receptor" to competitively bind to IL-33, reducing myocardial protection, leading to myocardial remodeling and dysfunction. Clinically, heart failure is diagnosed and the prognosis is evaluated by detecting the ST2 content in the blood. The normal population is ≤35ng / ml, and the patients with poor prognosis of heart failure are >35ng / ml. In addition, the detection of ST2 helps to assess the risk of acute coronary syndrome (ACS). High levels of ST2 are associated with a higher incidence of cardiovascular events.

[0003] Currently, the main methods for detecting ST2 in clinical practice include chemiluminescence, immunochromatography and latex turbidimetry. These methodologies are all based on the principle of double antibody sandwich method for antigen detection. Therefore, it is crucial to obtain monoclonal paired antibodies for ST2 with high affinity and strong specificity. This is also a key factor limiting the clinical detection of ST2. Summary of the invention

[0004] In order to overcome the shortcomings of the prior art, the present invention has developed ST2 monoclonal paired antibodies that can be used for clinical detection, which are used for detection and capture respectively; in addition, the present invention also provides a method for clinical detection of ST2 antigen, which can quantitatively detect the content of ST2 in blood.

[0005] In order to solve the above problems, the technical solution provided by the present invention is as follows:

[0006] A monoclonal antibody against human ST2, wherein the monoclonal antibody is 1D2, the amino acid sequence of the heavy chain variable region of the 1D2 antibody is SEQ ID NO: 2, and the amino acid sequence of the light chain variable region is SEQ ID NO: 3.

[0007] A monoclonal antibody against human ST2, wherein the monoclonal antibody is 5C6, the amino acid sequence of the heavy chain variable region of the 5C6 antibody is SEQ ID NO:4, and the amino acid sequence of the light chain variable region is SEQ ID NO:5.

[0008] A monoclonal antibody against human ST2, wherein the monoclonal antibody is 1C3, the amino acid sequence of the heavy chain variable region of the 1C3 antibody is SEQ ID NO: 6, and the amino acid sequence of the light chain variable region is SEQ ID NO: 7.

[0009] A monoclonal antibody against human ST2, wherein the monoclonal antibody is 120, the amino acid sequence of the heavy chain variable region of the 120 antibody is SEQ ID NO: 8, and the amino acid sequence of the light chain variable region is SEQ ID NO: 9.

[0010] A monoclonal antibody against human ST2, wherein the monoclonal antibody is 120-1, the amino acid sequence of the heavy chain variable region of the 120-1 antibody is SEQ ID NO: 10, and the amino acid sequence of the light chain variable region is SEQ ID NO: 11.

[0011] The present invention develops ST2 monoclonal paired antibodies that can be used for clinical detection, which are used for detection and capture respectively; the paired antibodies are obtained through mouse hybridoma cell screening technology, which solves one or more of the above-mentioned needs or limitations, and provides a methodology that can be used for clinical sample detection - time-resolved fluorescent microsphere immunochromatography, which is used to quantitatively detect the content of ST2 in blood.

[0012] The method comprises the following steps:

[0013] 1) Recombinant expression of ST2: The eukaryotic expression plasmid of ST2 was obtained by molecular cloning technology, recombinant expression was performed in 293 cells, and high-purity recombinant ST2 protein was obtained by affinity purification.

[0014] 2) Hybridoma cell screening: The recombinant ST2 protein is subjected to four rounds of immunization. The spleens of the immunized mice are removed to isolate B cells. The isolated B cells are fused with myeloma cells, and positive clones are screened out through pressure screening, limiting dilution, ELISA detection and other methods. Finally, the heavy and light chain sequences of the antibody are obtained through molecular biological methods.

[0015] 3) Expression of ST2 antibody: Gene synthesis of ST2 heavy and light chain sequences and codon optimization were performed, and constructed into eukaryotic expression vectors, such as transient vectors such as pCDNA3.1, pCDNA, PTT5, etc., and stable expression vectors with resistance genes can also be selected. In this embodiment, the vector was constructed into pCDNA3.4 containing mouse IgG constant region for transient expression in 293 cells;

[0016] 4) ST2 antibody purification: affinity purification of ST2 antibody was performed using protein A filler, the eluent was 0.1-0.2 M glycine (pH = 2.5-3.0), and finally neutralized with 1-2 M Tris-HCl (pH = 8.0-8.5). The dialysate was selected from Tris-HCl buffer or PBS buffer, with a suitable pH of 6.0-8.0 and an optimal pH of 7.0;

[0017] 5) ELISA method to detect the affinity of ST2 antibody: 500ng / ml-2000ng / ml recombinantly expressed ST2 protein was coated, 100ul per well, 500ng / ml in the embodiment of the present invention, coated at 4°C for 20hr, and washed 5 times with PBST on the next day; 100μg / ml ST2 antibody was added and diluted in 3-fold gradient to 15 wells, and negative wells were set, i.e., the antibody was replaced by the diluent, and incubated at 37°C for 2 hours.

[0018] 6) SPR detection of affinity and pairing: SPR detection of affinity and pairing:

[0019] Based on the principle of surface plasmon resonance (SPR), affinity and pairing detection were performed. ST2 antigen was diluted to 30ug / ml, 50ul was aspirated into the corresponding well of Ni NTA chip, the lid was closed, and the start button was clicked to fix the ligand for 5min. After the fixation procedure was completed, the ST2 sample was shaken off and patted dry. The diluted antibody was added to the target chip wells at the same time according to the concentration gradient with a discharge gun, 50μL / well, the lid was closed, and then the software start button was quickly clicked to perform the binding kinetics detection process of the analyte and the ligand for 5min. After the end, the antibody sample was shaken off and patted dry. 150ul of dilution buffer was added to the experimental well, the lid was closed, and the start button was clicked to perform the analyte dissociation process for 10min. Finally, data statistics and analysis were performed.

[0020] The same method can be used for paired detection of ST2 antibodies. Dilute the ST2 antigen to 20ug / ml, aspirate 50ul into the corresponding hole of the Ni NTA chip, cover the lid, and click the start button to fix the ligand for 5 minutes. After the fixation program is completed, shake off the ST2 sample, pat dry, add 50ul of the diluted first ST2 antibody, close the lid, and quickly click the software start button to detect the binding of the analyte and the ligand for 5 minutes. After the end, shake off the antibody sample, pat dry, add 50ul of the diluted second ST2 antibody, close the lid, and quickly click the software start button to detect the binding of the analyte and the ligand for 5 minutes. After the end, shake off the antibody sample, pat dry, add 50ul of the diluted second ST2 antibody, close the lid, and quickly click the software start button to detect the binding of the analyte and the ligand for 5 minutes. After the end, shake off the antibody sample, and finally perform paired data analysis.

[0021] 7) Time-resolved microsphere immunochromatography detection: The capture antibody and rabbit anti-chicken IgY were coated on the NC membrane, with the capture antibody as the T line and the rabbit anti-chicken IgY as the C line, and dried at 45°C overnight; the detection antibody was coupled to the time-resolved microspheres, 1mg of the time-resolved microspheres was coupled to 0.1mg of the antibody, and after the coupling was completed, the microspheres were sprayed onto the binding pad and dried at 45°C overnight; the NC membrane, gold pad, sample pad, and absorbent paper were assembled and cut into strips and loaded into the immunochromatography cartridge for testing of blank solution and antigen solution, the blank solution and antigen solution were added in a volume of 80ul, the detection time was 10min, and finally data analysis and statistics were performed.

[0022] The beneficial effects of the present invention are as follows: the present invention develops ST2 monoclonal paired antibodies that can be used for clinical detection, which are used for detection and capture respectively; the paired antibodies are obtained by mouse hybridoma cell screening technology, and a methodology that can be used for clinical sample detection-time-resolved fluorescent microsphere immunochromatography is provided, which is used for quantitative detection of the content of ST2 in the blood. ST2 recombinant protein is obtained by eukaryotic cells and can be used as a clinical detection standard. Paired antibodies with high affinity and strong specificity are obtained by mouse hybridoma cell technology, which are used for detection and capture respectively, and can be used for the detection of clinical samples. A method that can be used for clinical sample detection-time-resolved microsphere immunochromatography detection is provided, which can be used for quantitative detection of ST2 protein in clinical samples, with a correlation coefficient>0.99 and a test range of 1-250ng / ml. The paired antibodies are used to detect clinical samples 10 times, and the CV is less than 15%, and the stability is good. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 It is a graph of serum titer ELISA assay data;

[0024] Figure 2 This is the SDS-PAGE electrophoresis diagram of the recombinantly expressed antibodies of 5 hybridoma cells;

[0025] Figure 3This is a graph showing the ELISA activity assay of five hybridoma cells;

[0026] Figure 4 This is the affinity graph of 5 antibodies detected by SPR;

[0027] Figure 5 This is a paired test diagram of 5 antibody strains;

[0028] Figure 6 It is a time-resolved microsphere immunochromatography assay to detect ST2 antigen. DETAILED DESCRIPTION

[0029] The present invention is further described in detail below with reference to the embodiments, but is not limited thereto.

[0030] Embodiment 1:

[0031] 1) Expression of ST2 antigen:

[0032] The ST2 antigen sequence was synthesized into the pCDNA3.4 eukaryotic expression vector, and the amino acid sequence was shown in SEQ NO: 1. The plasmid was transformed into the DH5a competent medium for bacterial preservation. The preserved glycerol bacteria were cultured at 180 rpm overnight, and the bacteria were collected by centrifugation at 8000 rpm for 10 min for plasmid extraction. The cells were inoculated with 1-1.5×10^6 / ml cells one day before transfection, and cultured at 37°C, 125 rpm, and 5% CO2. Cell counts were performed before transfection, and the cell number was 2.5-3×10^6 / ml. 6 / ml, transfection can be performed when the viability is ≥96%; transfection is performed using PEI max transfection reagent, and 2ml of feed solution is added 20hr after transfection; centrifugation is performed at 8000rpm for 6min after 4 days of expression, and the supernatant is collected for purification; the precipitate is discarded after cell centrifugation, and the supernatant is taken. 100ml of the culture medium supernatant is directly purified with 1ml Ni Smart Beads, and the filler is loaded into a gravity column tube; 20 times the column volume of balance solution is used for balance, and the balance solution can be PBS buffer or Tris buffer; the sample is loaded after the balance is completed; after the loading is completed, the sample is rinsed again with 20 times the column volume of balance solution; the elution is collected in separate tubes, and the eluent is 250mM imidazole, one column volume is collected in each tube, and a total of 5 tubes are collected; the sample is dialyzed into PBS solution overnight.

[0033] 2) ST2 antibody hybridoma cell screening

[0034] The recombinantly expressed ST2 antigen was used to immunize mice. The antigen was diluted with PBS and then mixed with the adjuvant in a volume ratio of 1:1. The BALB / c mice were subcutaneously injected with a dose of 0.2 mL. Two weeks after the first immunization, a booster immunization was performed, and a total of 4 booster immunizations were performed. Finally, blood was collected from the tail vein for titer determination. ELISA used 100 ng of ST2 recombinant antigen for overnight coating at 4°C; after discarding the coating solution, the cells were blocked at 37°C for 1 hour and washed 5 times with PBST, and then 100 μl of mouse serum with different dilution gradients was added; after washing 5 times with PBST, a 1:5000 diluted goat anti-rabbit secondary antibody was added, 100 μl per well, and incubated at 37°C for 1 hour; after washing 5 times with PBST, 100 ul TMB was directly added for color development, and finally 0.25 M HCl was added to terminate the reaction. The results are shown in Figure 1 After the serum was diluted 1:102400, the OD450 was around 1.2, indicating that the antibody titer was qualified.

[0035] Take out the spleen of the immune mouse to be fused in the clean bench, transfer it to the cell screen, grind it and dilute it with serum-free medium, then place the cells in the incubator for preheating and activation. Also take out the thymus of the healthy mouse in the clean bench, transfer it to the prepared cell screen, grind it and resuspend the cell pellet with 1ml serum, and set aside.

[0036] Prepare PEG and serum-free medium and preheat them in the incubator. Take out the prepared SP2 / 0 and spleen cells from the incubator and mix them in a 50ml centrifuge tube at a ratio of SP2 / 0: B cells = 1:5. Mix thoroughly. Centrifuge the mixed suspension and discard the supernatant. Add PEG for cell fusion. After fusion for 1 minute, add serum-free medium to terminate the reaction. Centrifuge again and discard the supernatant. Resuspend the precipitate with HAT medium and evenly spread the cells into a 96-well plate with a spray gun.

[0037] After 4-5 days of culturing the fused cells, the medium was changed, all the medium was sucked out with a spray gun, and then 20% HAT medium was added, and finally cultured in an incubator. When the cells in the well grew to about 80%, ELISA was performed, and finally the five hybridoma cell lines 1D2, 5C6, 1C3, 120 and 120-1 were confirmed to be active.

[0038] 3) ST2 antibody sequence analysis

[0039] The obtained 5 ELISA positive clones were gene retrieved by degenerate primers, and the retrieved sequences were all consistent with the characteristics of antibody sequences after analysis by Kabat database. The amino acid sequence of the heavy chain variable region of the 1D2 antibody is shown in SEQ ID NO:2, and the amino acid sequence of the light chain variable region is shown in SEQ ID NO:3; the amino acid sequence of the heavy chain variable region of the 5C6 antibody is shown in SEQ ID NO:4, and the amino acid sequence of the light chain variable region is shown in SEQ ID NO:5; the amino acid sequence of the heavy chain variable region of the 1C3 antibody is shown in SEQ ID NO:6, and the amino acid sequence of the light chain variable region is shown in SEQ ID NO:7; the amino acid sequence of the heavy chain variable region of the 120 antibody is shown in SEQ ID NO:8, and the amino acid sequence of the light chain variable region is shown in SEQ ID NO:9; the amino acid sequence of the heavy chain variable region of the 120-1 antibody is shown in SEQ ID NO:10, and the amino acid sequence of the light chain variable region is shown in SEQ ID NO:11.

[0040] 4) Recombinant expression and purification of ST2 antibody

[0041] The retrieved sequence was fully synthesized into the pCDNA3.4 vector containing mouse IgG1. The plasmid was transformed into DH5a competent medium for bacterial preservation. The stored glycerol bacteria were cultured at 180rpm overnight, and centrifuged at 8000rpm for 10min to collect the bacteria for plasmid extraction; the cells were inoculated with 1-1.5×10^6 / ml cells the day before transfection, and cultured at 37℃, 125rpm, and 5% CO2; cells were counted before transfection, and transfection was performed when the cell count reached 2.5-3×10^6 / ml and the viability was ≥96%; transfection was performed using PEImax transfection reagent, and 2ml of feed solution was added 20hr after transfection; after 4 days of expression, the supernatant was collected for purification at 8000rpm for 6min; the precipitate was discarded after cell centrifugation, and the supernatant was taken. 200ml of supernatant was purified with 2ml rProtein At; 2ml rProtein At was added to the supernatant. The column was loaded with At filler and balanced with 20 column volumes of balancing solution, which can be PBS buffer or Tris buffer; the sample was slowly loaded after the balancing; after the loading, the sample was continuously washed with 20 column volumes of balancing solution; after the balancing, the column was eluted and collected in separate tubes, the eluent was 0.1M Glycine (pH=3.0), and 2 ml of eluent was collected in each tube; after the elution, the pH was directly adjusted to neutral with 2M Tris (pH=8.0); the purity and concentration were detected by electrophoresis, and the column was dialyzed into 1×PBS (pH=7.4) overnight. The SDS-PAGE electrophoresis diagram of the three antibodies after expression is shown in FIG. Figure 2 From the figure, we can see that the non-reducing bands of the five antibodies are around 180KDa, the heavy chain of the reducing bands are around 55KDa, and the light chain is around 25KDa.

[0042] 5) ELISA to detect the affinity of 1D2, 5C6, 1C3, 120 and 120-1 antibodies

[0043] ELISA uses 100ng of ST2 recombinant protein for coating at 4℃ overnight; discard the coating solution and wash with PBST 5 times, then block at 37℃ for 1 hour, then add 100μl ST2 recombinant antibody with different dilution gradients, the starting concentration of 5 antibodies is 1000ng / ml, 100μl per well, then dilute five times, and set up a negative control well; after washing with PBST 5 times, add 1:5000 diluted goat anti-rabbit secondary antibody, 100μl per well, incubate at 37℃ for 1 hour; after washing with PBST 5 times, directly add 100ul TMB for color development, and finally add 0.25M HCl to terminate the reaction. The results are shown in Figure 3 , all five recombinant antibodies had high ELISA activity.

[0044] 6) SPR detection of affinity of 5 antibodies

[0045] The affinity of 5 ST2 antibodies was tested using the SPR multifunctional detection device. The chip used was Ni NTA. First, the ST2 antigen was diluted to 30ug / ml, and 50ul was aspirated into the corresponding well of the Ni NTA chip. The lid was closed and the start button was clicked to fix the ligand for 5min. After the fixation procedure was completed, the ST2 sample was shaken off and patted dry. The diluted antibodies were added to the target chip wells at the same time according to the concentration gradient with a spray gun, 50μL / well, the lid was closed, and then the software start button was quickly clicked to detect the binding kinetics of the analyte and the ligand for 5min. After the end, the antibody sample was shaken off and patted dry. 150ul of dilution buffer was added to the experimental well, the lid was closed, and the start button was clicked to perform the analyte dissociation process for 10min. Finally, data statistics and analysis were performed. The results are shown in the figure. Figure 4 , the affinity of 1C3 is 4×10^-9M, the affinity of 1D2 is 4×10^-7M, the affinity of 5C6 is 3×10^-7M, the HM120 KD=3.6810^-10M, and the affinity of HM120-1 is 1.6×10^-9M.

[0046] 7) SPR detection of 5 antibody pairs

[0047] Dilute ST2 antigen to 20ug / ml, pipette 50ul into the corresponding hole of Ni NTA chip, cover the lid, click the start button to fix the ligand for 5min, after the fixation program is finished, shake off the ST2 sample, pat dry, add 50ul of the diluted first ST2 antibody, close the lid, and quickly click the software start button to detect the binding of analyte and ligand for 5min, after the end, shake off the antibody sample, pat dry, add 50ul of the diluted second ST2 antibody, close the lid, and quickly click the software start button to detect the binding of analyte and ligand for 5min, after the end, shake off the antibody sample, and finally perform paired data analysis. The results are shown in Figure 5 , 1C3, HM120-1 and 5C6 showed different epitopes.

[0048] 8) Time-resolved microsphere immunochromatography to detect ST2 antigen

[0049] Commercial ST2 capture antibody and detection antibody were purchased, and the time-resolved microsphere immunochromatography was prepared in the same way. Specifically, the capture antibody and rabbit anti-chicken IgY were coated on the NC membrane, the detection antibody was used as the T line, and the rabbit anti-chicken IgY was used as the C line, and the samples were dried at 45°C overnight; the 1C3 antibody was sprayed with gold to capture and coupled to the time-resolved microspheres, 1mg of the time-resolved microspheres was coupled to 0.1mg of the antibody, and after the coupling, the microspheres were sprayed onto the binding pads, and the samples were dried at 45°C overnight; the NC membrane, gold pad, sample pad, and absorbent paper were assembled and cut into strips and loaded into the immunochromatography cartridge to test the blank solution and antigen solution. The blank solution and antigen solution were added in a volume of 80ul, and the detection time was 10min. Finally, the T value / C value was statistically analyzed, and the results are shown in Figure 6 , HM120-1 and 1C3 were used as paired antibodies for ST2 detection, and the correlation was 0.9977, with a detection range of 1-250ng / ml; while the correlation of commercial ST2 paired antibodies for ST2 detection was 0.9943, and the detection limit was lower than that of HM120-1 and 1C3, indicating that the antibodies of the present invention have higher sensitivity and are superior to commercial antibodies.

[0050] The above is only a preferred embodiment of the present invention. It should be pointed out that a person skilled in the art can make several modifications and improvements without departing from the inventive concept of the present invention, which all belong to the protection scope of the present invention.

[0051] Sequence Listing

[0052] SEQ ID NO: 1

[0053] KFSKQSWGLENEALIVRCPRQGKPSYTVDWYYSQTNKSIPTQERNRVFASGQLLKFLPAAVADSGIYTCIVRSPTFNRTGYANVTIYKKQSDCNVPDYLMYSTVSGSEKNSKIYCPTIDLYNWTAPLEWFKNCQALQGSRYRAHKSFLVIDNVMTEDAGDYTCKFIHNENGANYSVTATRSFTVKDEQGFSLFPVIGAPAQNEIKEVEIGKNANLTCSACFGKGTQFLAAVLWQLNGTKITDFGEPRIQQEEGQNQSFSNGLACLDMVLRIADVKEEDLLLQYDCLALNLHGLRRHTVRLSRKNP

[0054] IDHHS

[0055] SEQ ID NO:2

[0056] EVQLQQSGAELVKPGASVKLSCTVSGFNIKDTYIHWVKQRPEQGLEWIGRIDPANDNTKYDPKFQDKATISSDTSSNAAFLQLNSLTSEDTAVYYCADGSPFNYFSLDYWGQGTSVTVSS

[0057] SEQ ID NO:3

[0058] DILMTQSTSSLSASLGDRVTISCRASQDISKYLNWYQQKADGTVKLLIYYTSRLDSGVPSRFSGSGSGTDYSLTITNLEQEDIATYFCQQGNTLPWTFGGGTKLELK

[0059] SEQ ID NO:4

[0060] QVQLQQSGPELKKPGETVKISCKTSGYTFTSSGINWVKQAPGKGLKWMGWINTYTRQPTYTADFKGRFAFSLETSASTAYLQINNLKNEDTATYFCATLPYYAMDYWGQGTSVTVSS

[0061] SEQ ID NO:5

[0062] DVVMTQTPLSLPVSLGDQASISCRSSQSLVHSNGNTYLHWYLQKPGQSPKLLIYKVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDLGVYFCSQSAHVPYTFGGGTKLEIK

[0063] SEQ ID NO:6

[0064] QVQLKQSGPGLVKPSQSLSLTCTVTGYSITSDYAWNWIRQFPGNKLEWVGFITYSAYTSYNPSLKSRISITRDTSKNQFFLQLNSVTTEDTATYYCARRYDYDFDYWGQGTTLTVSS

[0065] SEQ ID NO:7

[0066] DIVMSQSPSSLAVSAGEKVTMSCKSSQSLLNSRIRKNNLAWYQQKPGQSPKLLIYWASTRESGVPDRFTGSGSGTDFTLTISSVQAEDLAVYYCKQSYNLFTFGGGTKLELK

[0067] SEQ ID NO:8

[0068] EVKLMESGGGLVQPGGSLKLSCAASGFTFNIYTMSWVRQTPEKRLEWVAYINNGGGSTYYPDTVKGRFTISRDNAKNTLYLQMSSLKSEDTAIYYCARQVTYFDYWGQGTTVTVSS

[0069] SEQ ID NO:9

[0070] DVQITQTPASLAASVGETVTITCRASENIYYTLAWYQQKQGKSPQLLIYNANTLEDGVPSRFSGSGSGTQYSMKINSTQPEDTATYFCQQSYGVPWTFGGGTKLELK

[0071] SEQ ID NO:10

[0072] QVQLQQSGPELKKPGETVKISCKASGYTFTNYGMNWVKQAPGKGLKWMGWINTYTGEPTYADDFKGRFAFSLETSASTAYLQINNLKNEDMATYFCARDYGPYYFNYWGQGTTLTVSS

[0073] SEQ ID NO:11

[0074] DIQMTQSPSSLSASLGERVSLTCRASQDIGFYLSWLQQEPDGTIKRLIYATSSLDSGVPKRFSGSRSGSDYSLTISSLESEDFVDYYCLQYASSPPTFGGGTRLEIK

Claims

1. A monoclonal antibody against human ST2, characterized in that: The monoclonal antibody is 1D2, the amino acid sequence of the heavy chain variable region of the 1D2 antibody is SEQ ID NO:2, and the amino acid sequence of the light chain variable region is SEQ ID NO:

3.

2. A monoclonal antibody against human ST2, characterized in that: The monoclonal antibody is 5C6, the amino acid sequence of the heavy chain variable region of the 5C6 antibody is SEQ ID NO:4, and the amino acid sequence of the light chain variable region is SEQ ID NO:

5.

3. A monoclonal antibody against human ST2, characterized in that: The monoclonal antibody is 1C3, the amino acid sequence of the heavy chain variable region of the 1C3 antibody is SEQ ID NO:6, and the amino acid sequence of the light chain variable region is SEQ ID NO:

7.

4. A monoclonal antibody against human ST2, characterized in that: The monoclonal antibody is 120, the amino acid sequence of the heavy chain variable region of the 120 antibody is SEQ ID NO:8, and the amino acid sequence of the light chain variable region is SEQ ID NO:

9.

5. A monoclonal antibody against human ST2, characterized in that: The monoclonal antibody is 120-1, the amino acid sequence of the heavy chain variable region of the 120-1 antibody is SEQ ID NO:10, and the amino acid sequence of the light chain variable region is SEQ ID NO:

11.

6. A carrier, characterized in that A monoclonal antibody against human ST2 comprising any one of claims 1 to 5.

7. Use of the anti-human ST2 monoclonal antibody according to any one of claims 1 to 5 in detecting ST2 antigen.

8. Use of the anti-human ST2 monoclonal antibody according to any one of claims 1 to 5 in capturing ST2 antigen.

9. Use of the anti-human ST2 monoclonal antibody according to any one of claims 1 to 5 in the preparation of an ST2 antigen detection reagent.

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