CN109734805A - Anti- CK20 protein monoclonal antibody, cell line and its preparation method and application - Google Patents
Anti- CK20 protein monoclonal antibody, cell line and its preparation method and application Download PDFInfo
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Abstract
The present invention relates to field of biological detection, a kind of anti-CK20 protein monoclonal antibody is provided, the amino acid sequence of heavy chain and light chain variable region is amino acid sequence shown in SEQ ID NO.4 and SEQ ID NO.5 respectively.Inventor additionally provides the preparation method of anti-CK20 protein monoclonal antibody, chooses 411-424 albumen of people CK20 amino acid and is coupled with KLH, as immunogene.Inventor additionally provides the hybridoma cell line of one plant of anti-CK20 albumen of secretion, and the cell line is mouse hybridoma cell system 10F6B12A9, deposit number are as follows: CGMCC NO.16204.Anti- CK20 protein monoclonal antibody has high specific, sensibility, and the cell of CK20 albumen can be expressed with specific recognition, is suitable for immunology detection, especially immunohistochemistry and detects.
Description
Technical field
The present invention relates to field of biological detection, in particular to anti-CK20 protein monoclonal antibody, cell line and its preparation side
Method and application.
Background technique
Cytokeratin (Cytokeratin, CK) is distributed across the median fiber silk in ectodermal origin cell, is cell
One of component of skeleton.It is divided into 20 kinds of CK1~CK20 etc..Cytokeratin is distributed mainly in umbrella organisations' cell, but
The abilities to express such as marrow, blood, lymph node are weaker, and tumour cell can retain the germinal layer characteristic in its source, and cytokeratin is
Become the more sensitive and special marker of tumour cell, is used to detection malignant cell.Proteins cytokeratin 20
(Cytokeratin 20, CK20) is one kind of I type cytokeratin, relative molecular mass 48000, coding DNA overall length
18kb contains 8 exons, 7 intrones.
The synthesis of CK20 is come across first in the mucous epithelium of the 8th week embryo, after be distributed widely in goblet cell and villus
In cell.CK20 has stringent epithelial tissue specificity in CK series.In normal tissue, as mammary gland, smooth muscle, blood are thin
Born of the same parents, lymphocyte, hematopoietic cell CK20 expression are all feminine gender, and positive expression is detected in intestinal mucosa cells, gastric mucosa and imprisons
Door gland cell, duodenal mucosa, urinary system umbrella cells, epidermis Merkel cell.The expression of CK20 is in cell
Continuous expression is positive when life, canceration, metastases, in vitro culture etc. change.The positive tumour of CK20 expression is seen: colorectum
Cancer, sdenocarcinoma of stomach and undifferentiated carcinoma, the urinary tract transitional cell carcinoma, biliary tract gland cancer, pancreatic ductal cell gland cancer, cutaneous Merkel cell carcinoma
Deng.CK20 expression feminine gender is seen: mucinous breast carcinoma and the non-myxoadenocarcinoma of lobular adenocarcinoma, carcinoma of endometrium, ovary, nephrocyte
Cancer, adenocarcinoma of lung and squamous cell carcinoma, squamous cell carcinoma (cervix, pharynx, esophagus, skin), sarcoma, malignant lymphoma etc..CK20
Characteristic distributions become good tumor markers, also be used to be authenticated tumor tissues source.
Preparing Monoclonal anti-CK20 antibody can be used to study CK20 albumen, and can be detected in tissue and cell using it
The expression of CK20 comes generation or migration progress antidiastole to tumour.But the CK20 monoclonal antibody of current high quality depends on state mostly
Outer import, testing cost is higher, and the background information that monoclonal antibody obtains is unintelligible.Therefore, this research prokaryotic expression people CK20 albumen,
Immune mouse, prepares CK20 monoclonal antibody, and identified, it is desirable to provide the state that a kind of background information is clear, property is stable
Productionization monoclonal antibody, detection and research for CK20.
Summary of the invention
Inventor provide a kind of anti-CK20 protein monoclonal antibody, the monoclonal antibody heavy and light chain variable region
Amino acid sequence is amino acid sequence shown in SEQ ID NO.4 and SEQ ID NO.5 respectively.
Further, the monoclonal antibody heavy and chain variable region amino acid sequence be respectively SEQ ID NO.2 and
Coded by nucleotide sequence shown in SEQ ID NO.3.
Further, the monoclonal antibody specificity identifies CK20 albumen.
Further, amino acid sequence shown in SEQ ID NO.1 in the monoclonal antibody specificity identification.
Further, the monoclonal antibody is generated by the hybridoma cell line that deposit number is CGMCC NO.16204.Institute
State cell strain be mouse hybridoma cell system 10F6B12A9, classification naming are as follows: mouse hybridoma cell system, the cell line in
In China Committee for Culture Collection of Microorganisms's common micro-organisms center's preservation, address is court, Beijing on 07 30th, 2018
No. 3 Institute of Microorganism, Academia Sinica, institute of positive area's North Star West Road 1.
Further, the anti-CK20 albumen is mouse IgG2aHypotype monoclonal antibody.
Inventor additionally provides a kind of preparation method of anti-CK20 protein monoclonal antibody, chooses CK20 Argine Monohydrochloride sequence
The 411st is arranged to the 424th amino acids and with carrier protein KLH as immunogene.
Inventor additionally provides the hybridoma cell line of one plant of anti-CK20 albumen of secretion, and the cell strain is Mouse Hybridoma Cells
Cell line 10F6B12A9, the cell line are preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, protect
Hiding number are as follows: CGMCC NO.16204.
Inventor also provides any of the above-described anti-CK20 protein monoclonal antibody, in the detection of CK20 protein immunization
Purposes.
Further, the immune detection includes immunohistochemical method, Western blot and enzyme-linked immunization.
It is different from the prior art, structure, antigenicity, group ammonification of the above-mentioned technical proposal according to CK20 albumen in conjunction with DNA
The hydrophilic and hydrophobic and secondary structure of base acid, selection, which has, is different from other similar albumen, suitable length and with special antigen
411-424 albumen of amino acid sequence (SEQ ID No.1) of property, region is as Antigenic Peptide.Sulfydryl modification is added simultaneously in N-terminal
It is coupled KLH, mouse is immunized as immunogen, through cell fusion, screening and subclone, obtains the anti-CK20 egg of efficient secretion
The monoclonal cell system 10F6B12A9 of white monoclonal antibody, and the anti-CK20 protein monoclonal as secreted by the cell line resist
Body.The antibody that this programme obtains has high specific, sensibility, and the cell of CK20 albumen can be expressed with specific recognition, is applicable in
It is detected in immunology detection, especially immunohistochemistry.
Detailed description of the invention
Fig. 1 is the polyacrylamide gel electrophoresis figure of CK20 monoclonal antibody after purification.
Fig. 2 is gastric tissue immunohistochemical staining result figure (CK20 that a left side is 10F6B12A9, the right side are commercially available CK20).
Fig. 3 is colonic tissue immunohistochemical staining result figure (CK20 that a left side is 10F6B12A9, the right side are commercially available CK20).
Fig. 4 is colon cancer tissue immunohistochemical staining result figure (CK20 that a left side is 10F6B12A9, the right side are commercially available CK20).
Specific embodiment
The preparation of 1 recombinant C K20 protein fragments of embodiment
One, gene cloning
From the CK20 protein sequence of selection number P35900 in Uniprot database (http://www.uniprot.org)
As standard sequence.According to its structure in conjunction with DNA, antigenicity, the hydrophilic and hydrophobic and secondary structure for forming amino acid, choosing
Select be different from other similar albumen, suitable length and have special antigenic region as Antigenic Peptide.Selected CK20 ammonia
Base acid sequence the 411st to the 424th bit sequence, corresponding nucleotides sequence is classified as SEQ ID No.1.Synthesize the above CK20 egg
It is white, its N-terminal addition sulfydryl modification and and carrier protein KLH, improve its immunogenicity, it is spare as immunogene.
The foundation of 2 10F6B12A9 hybridoma cell line of embodiment
One, it is immunized
CK20 albumen-KLH the conjugate obtained in embodiment 1 is diluted to 1mg/mL, then with it is isometric completely not
Family name's adjuvant (CFA, Sigma company) mixing and emulsifying carries out the Balb/c mouse (being purchased from Foochow Wu Shi experimental animal) of 18-20g
Abdomen injection is immune, and injection dosage is 50 μ g/.Hereafter every 14 days booster immunizations are primary, and antigen uses Freund non-fully adjuvant
(IFA, Sigma company) emulsification, dosage are 25 μ g/.It is examined with indirect ELISA (wavelength 450nm) within 14 days after 2nd booster immunization
The how anti-potency of anti-immunity original in mice serum is surveyed, the highest mouse of potency is immune with tail vein injection impact, and antigen is molten with PBS
Liquid mixes, and dosage is 50 μ g/.
Two, cell fusion
Mouse boosting cell suspension up to standard is immunized in sterile preparation, with murine myeloma cell sp2/0 (ATCC) with 5:1 ratio
Mixing, 1000rpm are centrifuged 10min, and after abandoning supernatant, the PEG (Sigma that 1mL is preheated to 37 DEG C is added from slow to fast in 1 minute
Company) solution, it needs gently to rotate centrifuge tube in adition process, comes into full contact with cell with PEG.After being stored at room temperature 90s, in 2min
Serum-free DMEM (Hyclone company) culture medium that 4mL is preheated to 37 DEG C is added from slow to fast, is added in subsequent 2min
10mL preheats plasma-free DMEM medium, adds remaining preheating plasma-free DMEM medium in last 2min, is settled to 50mL,
Entire adition process needs to slowly shake centrifuge tube, it is ensured that is uniformly mixed, mitigates the injury to cell.After being stored at room temperature 10min
It is centrifuged (1000rpm, 5min), abandons supernatant, cell is resuspended with 10-20mLHAT (Sigma company) culture medium, and with HAT culture medium
It is diluted to final concentration of 0.5 × 106Cells/mL, all solution are transferred in 96 orifice plates, 200 holes μ L/, and are marked.It will
96 well culture plates are carefully transferred to 37 DEG C, cultivate in 5%CO2 incubator.The growth conditions for inspecting periodically cell and potentially dirt
Dye, pays attention to opening and closing incubator less as far as possible, it is ensured that culture environment is stablized.The 5th day after fusion, HAT culture medium is added into culture plate,
50 holes μ L/.
Three, ELISA screens positive hybridoma cell
When fused cell diameter it is long to about 1-2mm when, draw 50-200 μ L culture solution supernatant and carry out cell sieve for the first time
Choosing (ELISA, IHC-P and other methods detection), while HAT culture medium is added to 200 μ L into culture hole.ELISA detection should
Culture solution supernatant, the culture hole inner cell culture solution that will test to obtain positive findings are fully transferred to 24 well culture plates, add HT
Culture medium, the hole 2mL/ are cultivated 3 days.
Repeat screening 24 orifice plates in each cell strain, reject be not positive findings culture hole cell, obtain positive findings compared with
Good culture hole cell.The positive hole cell that these 24 well culture plates obtain is subjected to subclone screening using limiting dilution assay,
Limiting dilution assay is obtained to be transferred to CO in cell liquid 96 well culture plates of addition2It is incubator culture 11 days, straight to clone cell
When diameter is 1-2mm, repeat cell screening.According to testing result, each subcloned cells strain, selection 4 are well-grown
Monoclonal positive culture hole, and be transferred in 24 orifice plates and continue to cultivate.Clone in 24 orifice plates is screened after a period of time again to obtain
Positive colony cell strain, as secrete the hybridoma cell strain 10F6B12A9 of specific monoclonal antibody.This cell strain is transferred to
It is expanded to logarithmic growth phase conservation in T-75 culture bottle or carries out subsequent experimental.
3 ascites of embodiment induces method preparation monoclonal antibody
One, prepared by ascites
It will be washed and be resuspended with serum free medium after cell strain culture to logarithmic growth phase, count 2 × 106A/mL.It is outstanding
10 days BALB/C mices of paraffin oil sensitization are used in floating cell solution intraperitoneal injection in advance, and every mouse injects 0.5mL.It raises small
Mouse started to acquire ascites after 7 days.The ascites of collection is centrifuged 10min in 4 DEG C, 8000rpm, draws supernatant solution and is collected in centrifuge tube
In, as ascites, 4 DEG C or -20 DEG C preservations.
Two, the purifying of monoclonal antibody
With rProtein A sepharose Fast Flow (GE company) affinity column antibody purification master from ascites
It is divided into: 1. fills column, the ProteinA filler of purchase is loaded in gravitational stratification column in right amount with equilibration buffer (0.1M Tris
Solution, pH7.0) it rinses to balance；2. loading will be added in the chromatographic column installed, control by the ascites of 0.22 μm of membrane filtration
1 drop/sec of flow velocity processed；3. balancing, Equilibration buffer wash to balance is used after upper complete sample liquid；4. eluting, elution buffer is added
(0.1M citric acid solution, pH4.5) rinses pillar and collects eluent；5. regenerating, equilibration buffer punching is added after the completion of elution
Pillar is washed to balancing, the 20% ethyl alcohol flushing of 2 times of column volumes is placed on 4 DEG C of preservations.
4 monoclonal antibody CHARACTERISTICS IDENTIFICATION of embodiment
One, subtype identification
Cell conditioned medium is diluted to 1 μ g/mL coated elisa plate, every hole adds 100 μ L, and 4 DEG C of coatings overnight, are emptied liquid, use
200 μ L confining liquids (the PBS-T solution containing 2%BSA) is added in PBS (PBS-T) containing 0.05%Tween board-washing 3 times, every hole, and 37
DEG C be incubated for 1h.It is emptied liquid, is cleaned 3 times with PBS-T.Every hole is added on the 0.1mL hybridoma cell strain culture solution for diluting 5 times
Clearly, 37 DEG C of incubation 1h.It is emptied liquid, is cleaned 3 times with PBS-T.With confining liquid 1:400 dilution HRP label sheep anti mouse (κ, λ,
IgM, IgG1, IgG2a, IgG2b, IgG3, IgA) and antibody (Southern Biotech company), every hole is separately added into 0.1mL, and 37 DEG C
It is incubated for 1h.It is emptied liquid, is cleaned 3 times with PBS-T.Every hole adds 100 μ LTMB (Huzhou Yingcheng Biological Technology Co., Ltd.) substrates
(the isometric mixed solution of A, B) develops the color, and reacts at room temperature 15min, it is anti-that 50 μ L 1N HCl solution color development stoppings are added in every hole
It answers, then the OD value under microplate reader measurement 450nm wavelength.The results show that monoclonal antibody of the present invention is IgG1Type source of mouse Dan Ke
Grand antibody.
Two, affinity costant measures
It is coated with CK20 albumen, peridium concentration is 100 μ g/ml, and 100 holes μ l/, overnight, PBS-T is washed 3 times 4 DEG C of coatings.Every hole
37 DEG C of 200 μ l confining liquid closing 1h, PBS-T is added to wash 3 times.The monoclonal antibody purified in embodiment 4 is diluted to following concentration
(unit: ng/mL): 2000,500,125,62.5,31.25,15.625,3.125,0.625,37 DEG C of incubations 1h, PBS-T wash 3
It is secondary.The sheep anti mouse secondary antibody 1:5000 dilution of HRP label, 100 μ l of every hole, 37 DEG C of incubation 1h, PBS-T are washed 3 times.100 μ are added in every hole
L TMB (Huzhou Yingcheng Biological Technology Co., Ltd.) developing solution, develop the color 13min, and 100 μ l 1.0N salting liquids is added to terminate reaction.With
The light absorption value of microplate reader measurement wavelength 450nm.The curve that OD value corresponds to antibody extension rate is drawn, 1/2 " platform OD value " is found out
Corresponding antibody concentration A.Calculating affinity costant using following equation is 7.16L × mol-1。
Three, monoclonal antibody atopic and application effect
Immunogen solution is selected, the identification specificity of monoclonal antibody of the invention is detected with the method for immunoblotting, it is right
CK20 albumen-KLH conjugate carries out 12% polyacrylamide gel electrophoresis.Gel protein is transferred to conventional wet robin
Pvdf membrane.Film is placed in 5%BSA-TBST solution (albumen is face-down), 1h is closed in 37 DEG C of shaking tables, to eliminate non-specificity
Background.5%BSA-TBST is washed off with TBST after closing, the monoclonal antibody of CK20 prepared by the present invention is added, in decoloration
Shaking table, which sways, is incubated for 1h.After washing film with TBST, secondary antibody is added, is incubated for 1h in decolorization swinging table, combines secondary antibody sufficiently with primary antibody.
Film is washed with TBST again, ECL colour reagent box, experimental result such as Fig. 1 is added, CK20 monoclonal antibody detects people
Shown in the western blot figure of CK20 albumen.As seen from Figure 1, band is single and color is deeper, illustrates the specificity of antibody and antigen
Significant reaction.
The dyeing of 6 organization chip of embodiment and identification
One, paraffin embedded tissues preparation process
HE slice dyeing is carried out to sample tissue, to determine lesion.It draws a circle to disease site, preparation punching.
When making acceptor wax block, plastics are placed on mold, the paraffin (fusing point is at 56~58 DEG C) of thawing are poured into mold, by tissue
Block adds appropriate wax liquor after being put into the wax liquor in mold be fully embedded in tissue block in wax liquor, by mould after being cooled to room temperature
Tool is put into -20 DEG C of refrigerator 6min, and wax stone is removed from the molds, and is sliced or is put into and saves backup in 4 DEG C of refrigerators.It is carried out after photo fix
Serial section, thickness are set to 4 μm, and serial section is floated in 40% alcohol, allows it to be unfolded naturally, then separated slice is transferred to
It opens up piece 30 seconds, is sliced with through the processed glass slide mount of 2%APES acetone solution, by manufactured organization chip in 45 DEG C of warm water
It is put into 60 DEG C of ovens and bakes piece 2 hours, take out room temperature cooling, be put into -4 DEG C of refrigerators and save.
Two, IHC dyeing and analysis
Conventional xylene dewaxes 3 times, and 6 minutes every time, aquation in 100%, 100%, 95%, 95%, 85% graded ethanol,
3 minutes every time, last tap water rinsed.Antigen retrieval is carried out, then slice is put into wet box, PBS is rinsed 3 × 3 minutes.Drop
Add 3%H2O2It is incubated for 10 minutes, PBS is rinsed 3 × 3 minutes.PBS is got rid of, peroxidase blocking reagent is added dropwise and is incubated at room temperature 10 points
Clock.Drying slice, is added dropwise the diluted primary antibody of proper proportion (diluting the dilution ratio according to antibody concentration come designerantibodies for the first time)
(25 DEG C) of room temperature are incubated for 1 hour, and PBS is rinsed 3 × 3 minutes, and secondary antibody is added dropwise and is incubated at room temperature 30 minutes, and PBS is rinsed 3 × 3 minutes, is got rid of
PBS is removed, is developed the color 3-10 minutes with the DAB developing solution of fresh configuration.Haematoxylin is redyed 20 seconds, and PBS returns indigo plant.According to 85% (3 points
Clock), 95% (3 minutes), 95% (3 minutes), 100% (3 minutes), 100% (3 minutes) alcohol gradient be successively dehydrated, finally
Transparent 10 minutes of dimethylbenzene twice, neutral gum mounting.
Immunohistochemical staining result is divided into: positive and negative.Positive expression must be in cell and the specific antigen portion of tissue
Position can just be considered as the positive.In tissue staining distribution clearly and in the accurate situation of cellular localization, coloration result is according to staining power
Difference carry out further division, it is specific as follows:
1, sample is weakly positive；Labeled as "+"；
2, sample is moderate positive；Labeled as " ++ "；
3, sample is High positive；Labeled as " +++ ".
4, sample is feminine gender, is labeled as "-".
Three, pattern detection result:
With anti-CK20 protein monoclonal antibody (10F6B12A9) and the commercially available anti-CK20 protein monoclonal antibody of control antibodies-
(mouse monoclonal antibody Ks20.8) synchronizes detection in 67 colorectal cancers, 58 gastric cancers, 53 oophoromas, 45 lung cancer, as a result
As shown in the table.
As the result is shown: the dyeing accurate positioning of anti-CK20 protein monoclonal antibody (10F6B12A9), dyeing is clearly and nothing but
Specific stain, clean background illustrate anti-CK20 protein monoclonal antibody (10F6B12A9) high specificity.Meanwhile resisting with commercially available
The positive of body CK20 (Ks20.8) is consistent with negative findings, illustrates this antibody in the specific with commercial antibody phase of tumor tissues
When.
And in some cases, being higher than using the positive staining intensity of anti-CK20 protein monoclonal antibody (10F6B12A9) is made
With contrast agents, illustrate that the sensibility of anti-CK20 protein monoclonal antibody (10F6B12A9) is higher than commercial antibody.
And it is anti-with anti-CK20 protein monoclonal antibody (10F6B12A9) and the commercially available anti-CK20 protein monoclonal of control antibodies-
Body (mouse monoclonal antibody Ks20.8) synchronizes detection, the sample positive and negative knot on the organization chip for including 30 kinds of normal tissues
Fruit is consistent, illustrates that this antibody is suitable with commercial antibody in the specificity of normal tissue.
30 kinds of normal tissues include: brain, heart, cerebellum, oesophagus, adrenal gland, stomach, ovary, small intestine, pancreas, Colon and rectum,
Parathyroid gland, liver, hypophysis, salivary gland, testis, kidney, thyroid gland, prostate, mammary gland, uterus, spleen, bladder, tonsillotome, bone
Flesh, thymus gland (child), skin, marrow, peripheral nerve, lung, mesothelial cell.
Fig. 2 is gastric tissue immunohistochemical staining result figure (CK20 that a left side is 10F6B12A9, the right side are commercially available CK20).
Fig. 3 is colonic tissue immunohistochemical staining result figure (CK20 that a left side is 10F6B12A9, the right side are commercially available CK20).
Fig. 4 is colon cancer tissue immunohistochemical staining result figure (CK20 that a left side is 10F6B12A9, the right side are commercially available CK20).
Sequence table
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Claims (10)
1. a kind of anti-CK20 protein monoclonal antibody, which is characterized in that the ammonia of the monoclonal antibody heavy and light chain variable region
Base acid sequence is amino acid sequence shown in SEQ ID NO.4 and SEQ ID NO.5 respectively.
2. monoclonal antibody according to claim 1, which is characterized in that the monoclonal antibody heavy and light chain variable region
Amino acid sequence is coded by nucleotide sequence shown in SEQ ID NO.2 and SEQ ID NO.3 respectively.
3. monoclonal antibody according to claim 1, which is characterized in that the monoclonal antibody specificity identifies CK20 egg
It is white.
4. monoclonal antibody according to claim 3, which is characterized in that the monoclonal antibody specificity identifies CK20 egg
Amino acid sequence shown in white middle SEQ ID NO.1.
5. a kind of anti-CK20 protein monoclonal antibody is generated by the hybridoma cell line that deposit number is CGMCC NO.16204.
6. monoclonal antibody according to claim 1, which is characterized in that the anti-CK20 albumen is mouse IgG2aHypotype list
Clonal antibody.
7. a kind of preparation method of anti-CK20 protein monoclonal antibody, it is characterised in that: choose CK20 protein amino acid sequence the
411 to the 424th amino acids and with carrier protein KLH as immunogene.
8. the hybridoma cell line of one plant of anti-CK20 protein monoclonal antibody of secretion, the cell line is mouse hybridoma cell system
10F6B12A9, the cell line are preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, deposit number
Are as follows: CGMCC NO.16204.
9. any anti-CK20 protein monoclonal antibody of claim 1-6, the purposes in the detection of CK20 protein immunization.
10. immune detection according to claim 9, which is characterized in that the immune detection includes immunohistochemical method,
Western blot and enzyme-linked immunization.
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| CN112390895A (en) * | 2020-12-11 | 2021-02-23 | 河南赛诺特生物技术有限公司 | CK20 antigen, hybridoma cell strain, monoclonal antibody and application thereof |
| CN113583120A (en) * | 2021-04-25 | 2021-11-02 | 福州迈新生物技术开发有限公司 | Monoclonal antibody of anti-CK 20 protein, cell strain, preparation method and application thereof |
| CN113817055A (en) * | 2021-10-27 | 2021-12-21 | 福州迈新生物技术开发有限公司 | anti-Actin protein monoclonal antibody, cell line and application thereof |
| CN113845592A (en) * | 2021-09-27 | 2021-12-28 | 福州迈新生物技术开发有限公司 | anti-CK 5/6 protein monoclonal antibody, cell strain thereof, preparation method and application |
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| CN112390895A (en) * | 2020-12-11 | 2021-02-23 | 河南赛诺特生物技术有限公司 | CK20 antigen, hybridoma cell strain, monoclonal antibody and application thereof |
| CN112390895B (en) * | 2020-12-11 | 2023-06-20 | 河南赛诺特生物技术有限公司 | CK20 antigen, hybridoma cell strain, monoclonal antibody and application thereof |
| CN113583120A (en) * | 2021-04-25 | 2021-11-02 | 福州迈新生物技术开发有限公司 | Monoclonal antibody of anti-CK 20 protein, cell strain, preparation method and application thereof |
| CN113583120B (en) * | 2021-04-25 | 2023-10-03 | 福州迈新生物技术开发有限公司 | Monoclonal antibody against CK20 protein, cell strain, preparation method and application thereof |
| CN113845592A (en) * | 2021-09-27 | 2021-12-28 | 福州迈新生物技术开发有限公司 | anti-CK 5/6 protein monoclonal antibody, cell strain thereof, preparation method and application |
| CN113845592B (en) * | 2021-09-27 | 2023-03-10 | 福州迈新生物技术开发有限公司 | anti-CK 5/6 protein monoclonal antibody, cell strain thereof, preparation method and application |
| CN113817055A (en) * | 2021-10-27 | 2021-12-21 | 福州迈新生物技术开发有限公司 | anti-Actin protein monoclonal antibody, cell line and application thereof |
| CN113845593A (en) * | 2021-10-27 | 2021-12-28 | 福州迈新生物技术开发有限公司 | anti-alpha-SMA protein monoclonal antibody, cell line and application thereof |
| CN113845593B (en) * | 2021-10-27 | 2023-03-10 | 福州迈新生物技术开发有限公司 | anti-alpha-SMA protein monoclonal antibody, cell line and application thereof |
| CN115028720A (en) * | 2022-06-28 | 2022-09-09 | 南京涵台余生物科技有限公司 | Exosomes and use of inhibitors to increase drug sensitivity in the treatment of cancer |
| CN115028720B (en) * | 2022-06-28 | 2023-09-05 | 成都赋智健康科技有限公司 | Exosomes and use of inhibitors to increase drug sensitivity in the treatment of cancer |
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