CN108070034B - Human soluble glycoprotein 130 monoclonal antibody, encoding gene, preparation method and application - Google Patents
Human soluble glycoprotein 130 monoclonal antibody, encoding gene, preparation method and application Download PDFInfo
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- CN108070034B CN108070034B CN201711353636.4A CN201711353636A CN108070034B CN 108070034 B CN108070034 B CN 108070034B CN 201711353636 A CN201711353636 A CN 201711353636A CN 108070034 B CN108070034 B CN 108070034B
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- monoclonal antibody
- sgp130
- protein
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- variable region
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Abstract
The invention discloses a base sequence and an amino acid sequence of a heavy chain variable region and a light chain variable region of a human soluble glycoprotein 130 monoclonal antibody, and also discloses a coding gene and application of the human soluble glycoprotein 130 monoclonal antibody, and a preparation method of the human soluble glycoprotein 130 monoclonal antibody. The human soluble glycoprotein 130 monoclonal antibody has the advantages of good specificity, high sensitivity and high purity on the result of Western-blotting, and can provide a relatively accurate detection means for the expression level of sgp130 and gp130 proteins in vivo.
Description
Technical Field
The invention belongs to the technical field of bioengineering, and particularly relates to a human soluble glycoprotein (sgp130) monoclonal antibody, a coding gene, a preparation method and application thereof
Background
gp130 is an important signal molecule, which is almost expressed on various tissue cells in vivo and is involved in some important physiological processes in vivo, and one of the main functions of gp130 is to utilize the extracellular region to bind to the IL-6-IL-6R complex and to transmit signals into cells. IL-6 is usually present in small amounts in human serum, but its content is greatly increased in the case of inflammation, infection, and tumor or cancer. In human serum, besides gp130, there is soluble form of gp130(sgp130), sgp130 is an extracellular domain fragment of gp130, and may exist in animals as an antagonist of the complex IL-6-IL-6R. The research shows that the content of sgp130 in the normal human plasma can reach 410 ng/ml. The sgp130 in vivo retains the ability to bind to ligands that block the anti-signaling pathway of IL-6, thereby reducing the incidence of inflammatory responses and, to a certain extent, inhibiting the development of tumors and certain cancers. The detection of the expression of the protein of sgp130 in vivo plays an important role in the research of drugs for treating tumors and cancers.
Disclosure of Invention
An object of the present invention is to provide a human soluble glycoprotein (sgp130) monoclonal antibody and a gene encoding the same.
The human soluble glycoprotein 130 monoclonal antibody comprises a light chain variable region and a heavy chain variable region, wherein the base sequence and the amino acid sequence of the heavy chain variable region are respectively shown as SEQ ID NO.1 and SEQ ID NO.2, and the base sequence and the amino acid sequence of the light chain variable region are respectively shown as SEQ ID NO.3 and SEQ ID NO. 4.
The human soluble glycoprotein 130 monoclonal antibody is a protein shown as the following A or B:
a: a protein in which the base sequence and the amino acid sequence of the heavy chain variable region are respectively composed of the sequences shown in SEQ ID NO.1 and SEQ ID NO.2, and the base sequence and the amino acid sequence of the light chain variable region are respectively composed of the sequences shown in SEQ ID NO.3 and SEQ ID NO. 4;
b: a-derived protein with the same function obtained by substituting and/or deleting and/or adding one or more bases/amino acid residues to the sequences shown in SEQ ID NO. 1-4.
The coding gene of the human soluble glycoprotein 130 monoclonal antibody is a DNA molecule shown in any one of the following 1) -3):
1) the nucleotide sequence of the heavy chain variable region is shown as SEQ ID NO.1, and the nucleotide sequence of the light chain variable region is shown as DNA molecule shown as SEQ ID NO. 3:
2) a DNA molecule which hybridizes to a DNA sequence defined in 1) and encodes an antibody according to claim 1 or 2;
3) a DNA molecule having 90% or more homology with the DNA molecule defined in 1) and encoding the antibody of claim 1 or 2.
The invention also provides a recombinant expression vector or an expression cassette or a transgenic cell line or a recombinant bacterium containing the human soluble glycoprotein 130 monoclonal antibody.
It is another object of the present invention to provide the use of a human soluble glycoprotein 130 monoclonal antibody for the preparation of a product that specifically binds to human soluble glycoprotein 130, or for the preparation of a medicament for the treatment of cancer, or for the detection of the presence of sgp130 in a sample.
The preparation method of the human soluble glycoprotein 130 monoclonal antibody adopts the following steps:
1) constructing recombinant plasmid of human soluble glycoprotein (sgp130) by means of molecular cloning, wherein the recombinant plasmid has the DNA molecular sequence of the coding gene of claim 4, transforming into host bacteria Rosseta (DE3) for protein expression, and purifying to obtain high-purity human soluble glycoprotein (sgp130) protein;
2) injecting prepared sgp130 antigen into Balb/c mouse body in abdominal cavity;
3) repeating the step 2) and boosting for 2 times, taking blood, and detecting the antibody titer in the serum by using an ELISA technology;
4) the titer of the antibody is taken to reach the requirement 106The spleen cells of the Balb/c mice are fused with myeloma cells to prepare hybridoma cells;
5) cloning and culturing hybridoma cells secreting the sgp130 monoclonal antibody for 2-3 times;
6) carrying out expanded culture on the hybridoma cells obtained by the cloning culture in the step 5) to obtain a monoclonal antibody of the anti-sgp 130 protein with higher concentration;
7) purifying the cell list antibody by using a Protein A affinity chromatography column;
8) and (3) specific detection: detecting the specificity of the obtained antibody by using a Western-blotting technology;
9) the heavy chain variable region of the prepared monoclonal antibody is subjected to gene sequencing by utilizing a PCR technology.
The method specifically comprises the following steps:
1) obtaining antigen sgp130 protein: the human sgp130 gene is located on human chromosome 5, the cDNA of sgp130 has a total length of about 1.9kb, is composed of 329 amino acids, and has a relative molecular mass (Mr) of about 60 kD. An upstream Primer and a downstream Primer of the sgp130 protein are designed by using software Primer 5.0, a gene of the sgp130 protein is obtained by amplification by using a Polymerase Chain Reaction (PCR) technology, the target gene and an expression vector pET-28a are subjected to double enzyme digestion respectively, then T4-DNA ligase is used for enzyme ligation, a recombinant plasmid pET-28a-sgp130 is obtained, and the gene sequencing is carried out. The recombinant plasmid pET-28a-sgp130 with correct sequencing is transformed into a host bacterium Rosseta (DE3) for protein expression. According to the results of polypropylene gel electrophoresis (SDS-PAGE), the sgp130 protein was expressed as inclusion bodies. The inclusion body of the sgp130 protein is dissolved in Tris-NaCl buffer solution containing 8M urea, and purified by using an artificial protein glue recovery kit, and the recovered sgp130 protein is dissolved in PBS buffer solution.
2) Diluting the sgp130 protein obtained in the step 1) to 1mg/mL by PBS, then mixing the sgp130 protein with an equal volume of complete Freund's adjuvant (IFC), emulsifying the mixture by an emulsifier until the mixture is not melted by dropping water, and selecting a Balb/c mouse with the age of 6-8 weeks for intraperitoneal immunization, wherein the injection dose is as follows: 100 ug/mouse.
3) Repeating step 2) two booster immunizations: on day 15, the Freund's incomplete adjuvant was changed to the injection dose: 50 mu g/mouse; day 29, freund's incomplete adjuvant, injected at a dose of: 50 ug/mouse.
4) 2 weeks after the third injection, tail blood was taken and serum titer was measured; ELISA is used for detecting the antibody titer in the blood serum of Balb/c mice, and the titer meets the requirement (higher than 10)6) The spleen cells of Balb/c mice are fused with myeloma cells to prepare hybridoma cells, the immunization is strengthened once three days before the fusion, no adjuvant is directly added, and the immunization dose is 50 mu g/mouse.
5) Fusing for about 2 weeks, screening the hybridoma cells by an ELISA method, taking myeloma cell culture supernatant as a negative control, taking immune mouse serum as a positive control, and taking the envelope antigen as sgp130 protein to obtain the hybridoma cells secreting the sgp130 protein monoclonal antibody.
6) Cloning and culturing the hybridoma obtained in the step 5) for 2-3 times, wherein each cloning and culturing time is about 7-10 days, observing the cells under an inverted microscope, marking holes which are visible by naked eyes and only grow single clone, and detecting the antibody; and performing amplification culture on the cells with the positive detection result again until 100% of pores secrete the sgp130 protein monoclonal antibody, establishing a line and storing the hybridoma cell strain.
7) And (3) carrying out amplification culture on the hybridoma cells obtained by cloning culture and screening in the step 6) to obtain the anti-sgp 130 protein high-concentration cell supernatant monoclonal antibody.
8) Mixing about 2g of Protein A filler with 10mL of Tris buffer solution with the pH value of 7.0, pouring the mixture into a chromatographic column, and standing for 5-10min to ensure that the filler is naturally settled to obtain a bubble-free Protein A filler column; adding 10 column volumes of Tris buffer pH 7.0 and washing the column at the appropriate flow rate; mixing the high-concentration monoclonal antibody obtained in the step 7) with a Tris buffer solution with the pH of 7.0 according to the volume ratio of 1:2, filtering the mixture by using a 0.45-micron filter membrane, adding the mixture into a chromatographic column for affinity chromatography, eluting the antibody by using an eluent (a pH 4.50.1M citric acid solution and a pH 8.0 neutralizing solution), dialyzing the eluent by using PBS for three times, and finally concentrating the antibody solution by using an ultrafiltration concentration tube to obtain the high-purity and high-concentration sgp130 monoclonal antibody.
9) And (3) carrying out gene sequencing on the light chain variable region and the heavy chain variable region of the prepared monoclonal antibody by utilizing a PCR technology.
The invention has the advantages that: the antibody is a completely humanized antibody, has high specificity, high affinity and high sensitivity, and is purified by Protein A filler to obtain monoclonal antibody with high purity. The monoclonal antibody prepared from the human-derived sgp130 protein can provide a more accurate detection means for the expression levels of the sgp130 and gp130 proteins in vivo.
Drawings
FIG. 1: agarose gel electrophoresis picture of sgp130 protein extracellular domain gene clone; wherein, M: marker DL 2000; 4: blank control; 1-3: PCR amplification product of sgp 130.
FIG. 2: a polyacrylamide gel electrophoresis chart for identifying the expression of the protein of the extracellular domain of the sgp130 protein; wherein M is Marker; 1: pET-28a-sgp130 induces the total protein of the pre-Rosseta; 2: total Rosseta protein 18h after induction with pET-28a-sgp 130; 3: after pET-28a-sgp130 is induced for 18h, crushing the supernatant; 4: pET-28a-sgp130 is induced for 18 hours and then the sediment is broken; circles are marked as target protein positions.
FIG. 3: WB result of detecting sgp130 protein by sgp130 monoclonal antibody.
Detailed Description
The following detailed description of specific embodiments of the present invention is provided in conjunction with examples, but it should be understood that the scope of the present invention is not limited to the specific embodiments.
The experimental procedures used in the following examples are all conventional procedures unless otherwise specified.
Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.
The human soluble glycoprotein 130 monoclonal antibody comprises a light chain variable region and a heavy chain variable region, wherein the base sequence and the amino acid sequence of the heavy chain variable region are respectively shown as SEQ ID NO.1 and SEQ ID NO.2, and the base sequence and the amino acid sequence of the light chain variable region are respectively shown as SEQ ID NO.3 and SEQ ID NO. 4.
The human soluble glycoprotein 130 monoclonal antibody is a protein shown as the following A or B:
a: a protein in which the base sequence and the amino acid sequence of the heavy chain variable region are respectively composed of the sequences shown in SEQ ID NO.1 and SEQ ID NO.2, and the base sequence and the amino acid sequence of the light chain variable region are respectively composed of the sequences shown in SEQ ID NO.3 and SEQ ID NO. 4;
b: a-derived protein with the same function obtained by substituting and/or deleting and/or adding one or more bases/amino acid residues to the sequences shown in SEQ ID NO. 1-4.
The coding gene of the human soluble glycoprotein 130 monoclonal antibody is a DNA molecule shown in any one of the following 1) -3):
1) the nucleotide sequence of the heavy chain variable region is shown as SEQ ID NO.1, and the nucleotide sequence of the light chain variable region is shown as DNA molecule shown as SEQ ID NO. 3:
2) a DNA molecule which hybridizes to a DNA sequence defined in 1) and encodes an antibody according to claim 1 or 2;
3) a DNA molecule having 90% or more homology with the DNA molecule defined in 1) and encoding the antibody of claim 1 or 2.
A human soluble glycoprotein 130 monoclonal antibody, for preparing a product specifically binding to human soluble glycoprotein 130, or for preparing a medicament for treating cancer, or for detecting whether a sample contains sgp 130.
Example 1
The preparation method of the human soluble glycoprotein 130 monoclonal antibody specifically comprises the following steps:
1) firstly, obtaining an antigen sgp130 protein: the sgp130 protein is selected as antigen, the number of bases is about 1878bp, and the molecular weight of the protein is about 60 kDa. Primer premier 5.0 is used for designing an upstream Primer of the extracellular domain of the sgp130 protein:
5 'GGAATTCCATATGCACCATCACCATCACCATGAGCTGCTGGACCCATGCG GCTATAT3' (plus His-Tag) (cleavage site EcoR I) and downstream primer:
5'CCGCTCGAGTCACACCAGCACAGGCACCACGATAGC 3' (restriction site, Xho I), amplifying by Polymerase Chain Reaction (PCR) technology to obtain a large amount of genes of the extracellular domain of sgp130 protein (shown in figure 1), carrying out double restriction on the target genes and a vector pET-28a respectively, then connecting by T4-DNA ligase to obtain a recombinant plasmid pET-28a-sgp130, and sending to gene sequencing. Transforming the recombinant plasmid pET-28a-sgp130 with correct sequencing into host bacteria Rosseta (DE3), picking out positive monoclonal colonies, inoculating the colonies into a 4mL test tube with 50 ug/mL Kanna resistant LB culture medium, and culturing at 37 ℃ and 200rpm/min overnight; inoculating 4mL of overnight culture into a triangular flask containing 50 mu g/mL of Carna resistant 100mL LB culture medium, culturing at 37 ℃ and 200rpm/min in a shaking way for 2-3 h until the OD value of the bacterial liquid reaches 1.0, adding isopropyl-beta-D-thiogalactoside (IPTG) to the final concentration of 1.0mM, changing the culture condition to 17 ℃, culturing at 200rpm/min for 16-18 h, collecting the bacterial bodies, suspending in 50mL of PBS, carrying out ultrasonic disruption, and carrying out polyacrylamide gel electrophoresis (SDS-PAGE) identification, wherein the result shows that: the sgp130 protein is expressed in the form of inclusion bodies. The inclusion body of the sgp130 protein is dissolved in Tris-NaCl buffer solution containing 8M urea, polyacrylamide gel electrophoresis is carried out (as shown in figure 2), and a protein gel recovery kit is used for recovery, so as to obtain the purified antigen protein.
2) Diluting the sgp130 protein obtained in step 1) to 1mg/mL with PBS, mixing with an equal volume of complete Freund's adjuvant (IFC), and emulsifying the mixture by using an emulsifier until the mixture is not changed by water drops; selecting Balb/c mice of 6-8 weeks old for abdominal multipoint injection immunization, wherein the injection dose of each mouse is 100 mu g, and taking blood three days before immunization as blank serum.
3) Repeating step 2) two booster immunizations: on day 15, the dose per mouse was 50 μ g; on day 29, each mouse was injected with 50 μ g of Freund's incomplete adjuvant for booster immunization.
4) 2 weeks after the third injection, tail blood was taken and serum titer was measured; detecting the antibody titer in Balb/c mouse serum by ELISA, taking blank control group serum as a negative control, and PBS as a blank control; the antibody titer is higher than 106Then, the spleen cells of the Balb/c mice are taken to be fused with myeloma cells to prepare hybridoma cells, which shows that the antibody has enough antibody titer and affinity.
TABLE 1 ELISA test of antibody titer in serum of BALb/C mice
5) Fusing for about 2 weeks, detecting and screening the antibody of the hybridoma cells by using an ELISA method, taking a hybridoma cell culture medium as a negative control, taking immune mouse serum as a positive control, and coating antigen of sgp130 protein with the concentration of 1 ug/ml; finally obtaining the hybridoma secreting the sgp130 protein monoclonal antibody.
6) Cloning, screening and culturing the hybridoma obtained in the step 5) for 2-3 times; culturing for 7-10 days, observing cells under an inverted microscope, marking holes which are only visible to the naked eye and are used for single clone growth, and detecting the antibody; and performing amplification culture on the cells with the positive detection result again until 100% of pores secrete the sgp130 monoclonal antibody, and establishing a line and storing the hybridoma cell strain.
Wherein, ELISA steps and results are as follows:
(1) coating: the sgp130 protein prepared according to the present invention was diluted to 1. mu.g/mL with coating buffer, and then coated on reaction wells of polystyrene plates at 100. mu.L/well overnight at 4 ℃.
(2) Washing: the coating solution in the wells was discarded, and the reaction wells were washed 3 times with wash buffer wells for 3X 3min (hereinafter referred to as washing).
(3) And (3) sealing: adding 200 mu L of sealing liquid into each hole, and sealing for 2h at 37 ℃; the confining liquid is discarded as much as possible.
(4) Adding a primary antibody: adding diluted cell culture supernatant of 5 times into each hole, and incubating for 1h at 37 ℃; discarding the primary antibody after incubation, fully washing and spin-drying.
(5) Adding a secondary antibody: adding 100 μ L of HRP-goat anti-mouse IgG secondary antibody diluted to a certain concentration with diluent into each well, and incubating for 30min at 37 ℃; and after incubation, discarding the enzyme-labeled secondary antibody, washing and spin-drying.
(6) Color development: adding 100 mu L of substrate TMB into each hole, and reacting for 15min at 37 ℃ in a dark place; 100mL of stop solution was added to each well to stop the color reaction. Determination of the absorbance at 450nm (A)450)。
TABLE 2 ELISA test fusion cell supernatant antibody titer table
And (3) analyzing an experimental result: from the ELISA results, it was found that the negative result (NC) was only 0.0515 and the positive result (PC) was OVER. The table marks the fusion cell lines with higher antibody titers, which are: a7, B2, B3, C7, C9, E9; these 6 cell lines had high specificity for sgp130 and produced many antibodies against sgp 130. These 5 cell lines were further subjected to cloning culture until a fused cell line with higher specificity and 100% secretion of monoclonal antibody was obtained.
7) Preparing the monoclonal antibody by a mouse ascites method: firstly, injecting 0.5mL of liquid paraffin into the abdominal cavity of each Balb/c mouse; after 10 days, 1X 10 injections are injected into the abdominal cavity of each Balb/c mouse6Step 6) dilution of serum-free DMEM medium with cell volume of 0.5mL, cloning culture and screeningSelecting the obtained hybridoma cells; observing Balb/c mice 7 days later, and collecting ascites if the abdomen of the mice is obviously enlarged; the ascites is first centrifuged at low speed to remove impurities such as cells, the supernatant is collected and then centrifuged at high speed to remove fine particles.
8) Mixing about 2g of Protein A filler with 10mL of Tris buffer solution with the pH value of 7.0, pouring the mixture into a chromatographic column, and standing for 5-10min to ensure that the filler is naturally settled to obtain a bubble-free Protein A filler column; adding 10 column volumes of Tris buffer pH 7.0 and washing the column at the appropriate flow rate; mixing the high-concentration sgp130 monoclonal antibody obtained in the step 7) with a Tris buffer solution with the pH value of 7.0 according to the volume ratio of 1:2, filtering by using a filter membrane with the diameter of 0.45 mu M, pouring into a chromatographic column for affinity chromatography purification, eluting non-specifically adsorbed impurities by using a Tris buffer solution with the pH value of 7.0 and the volume of 10 times of the column volume, eluting sgp130 monomer by using an eluent (pH 4.50.1M citric acid solution plus a neutralizing solution with the pH value of 8.0), dialyzing the eluent by using PBS for three times, and finally concentrating an antibody solution by using an ultrafiltration concentration tube to obtain the sgp130 monoclonal antibody with high purity and high concentration.
9) specific detection of sgp130 monoclonal antibody: detection is carried out by using Western-blotting, SDS-PAGE electrophoresis is carried out on the sgp130, a film of a target protein position is cut according to a Marker band, and the detection result comprises the steps of film transfer, primary antibody incubation, secondary antibody incubation, ECL color development and the like, so that the sgp130 monoclonal antibody disclosed by the invention has good specificity and can be used for well detecting a sample containing the sgp130 protein (as shown in figure 3).
10) Assay of sgp130 monoclonal antibody subtype: the subtype identification kit of the mouse monoclonal antibody is utilized, the ELISA detection method is adopted to carry out the subtype identification on the sgp130 monoclonal antibody purified by the ascites of the Balb/c mouse, and the result shows that: the sgp130 monoclonal antibody belongs to the subclass IgG1, and its light chain is a kappa chain.
TABLE 3 ELISA results for sgp130 monoclonal antibody subtypes
And (3) analyzing an experimental result: according to the ELISA result table of antibody subtype identification, the results of IgG1 and kappa are all OVER, so that the antibody heavy chain subtype of the invention is IgG 1; the antibody light chain subtype is kappa chain.
11) The heavy chain variable region and the light chain variable region of the prepared monoclonal antibody are subjected to gene sequencing by utilizing a PCR technology.
Example 2
The detection of the sgp130 protein by using a Western-blotting technology comprises the following specific operation steps:
1) biological samples containing the sgp130 protein were separated by 15% polyacrylamide gel electrophoresis (SDS-PAGE), and after completion of the electrophoresis, PAGE was removed, and the resulting gel was sandwiched with PVDF membrane, followed by wet-transfer.
2) After the completion of the electrotransfer, the PVDF membrane was removed, 5% skim milk powder-TBST (protein side down) was added, and the mixture was sealed for 1 hour at 37 ℃ with shaking (65rpm) in a shaker to eliminate the nonspecific background.
3) After the blocking is finished, 5% of skimmed milk powder-TBST is washed away by TBST, and primary antibody is added: the monoclonal antibody of sgp130 prepared by the invention is incubated for 1h (60rpm) in a decoloration shaker or for 12-16h overnight (4 ℃) to ensure that the primary antibody is combined with the specific protein.
4) Primary antibody was recovered and washed 4 times (60rpm) with TBST on a shaker for 5min each.
5) Adding the secondary antibody, and incubating for 1h in a decoloration shaker (60rpm) to ensure that the secondary antibody is fully combined with the primary antibody.
6) The secondary antibody was recovered and washed 4 times (60rpm) with TBST on a shaker for 5min each
7) Incubate for 3min with ECL color kit (200. mu.l reagent A/plate + 200. mu.l reagent B/plate). Removing the reaction solution, transferring to a preservative film, tabletting, developing and fixing. After fixation, the solution is naturally dried, a Canon camera shoots the solution, and the experimental results are recorded (as in 3).
While the invention has been described in further detail with reference to specific preferred embodiments thereof, it will be understood by those skilled in the art that the invention is not limited to the details of construction and that various changes in form and details may be made therein without departing from the spirit and scope of the invention.
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Claims (4)
1. A human soluble glycoprotein 130 monoclonal antibody characterized by: the antibody comprises a light chain variable region and a heavy chain variable region, wherein the base sequence and the amino acid sequence of the heavy chain variable region are respectively shown as SEQ ID NO.1 and SEQ ID NO.2, and the base sequence and the amino acid sequence of the light chain variable region are respectively shown as SEQ ID NO.3 and SEQ ID number 4.
2. The gene encoding the human soluble glycoprotein 130 monoclonal antibody of claim 1.
3. The encoding gene of claim 2, wherein: the coding gene is a DNA molecule shown as follows: the nucleotide sequence of the heavy chain variable region is shown as SEQ ID NO.1, and the nucleotide sequence of the light chain variable region is shown as DNA molecule shown as SEQ ID NO. 3.
4. The use of the monoclonal antibody against human soluble glycoprotein 130 of claim 1, wherein: the antibody is used for preparing products which specifically bind to the human soluble glycoprotein 130.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| WO2004113383A3 (en) * | 2003-06-23 | 2005-06-23 | Conaris Res Inst Ag | Pegylated soluble gp130-dimers useful as a medicament |
| CN104744582A (en) * | 2015-04-03 | 2015-07-01 | 福州大学 | gp130 extracellular domain protein as well as production method and application thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2004113383A3 (en) * | 2003-06-23 | 2005-06-23 | Conaris Res Inst Ag | Pegylated soluble gp130-dimers useful as a medicament |
| CN104744582A (en) * | 2015-04-03 | 2015-07-01 | 福州大学 | gp130 extracellular domain protein as well as production method and application thereof |
Non-Patent Citations (1)
| Title |
|---|
| 抗gp130单克隆抗体的研制及对IL-6 R信号的调控性初探;苗平 等;《第八届全国免疫学学术大会论文集》;20121231;全文 * |
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