CN101942429A - Purification method for recombined human tissue type plasminogen exciter TNK mutant rhTNK-tPA - Google Patents
Purification method for recombined human tissue type plasminogen exciter TNK mutant rhTNK-tPA Download PDFInfo
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Abstract
The invention relates to a purification method for recombined human tissue type plasminogen exciter TNK mutant rhTNK-tPA. The method comprises the following steps of: 1, filtering the rhTNK-tPA cell culture medium and taking the supernatant, and after chromatography by a BlueSepharose6FastFlow compatible column, eluting by using eluent a to obtain eluent a1 containing a target peak; 2, after performing chromatography of the eluent a1 containing the target peak obtained in step 1 by a LysineHyperD compatible column, eluting by using eluent b to obtain eluent b1 containing the target peak; and 3, performing SephadexG-25 gel filtration of the eluent b1 containing the target peak obtained in step 2, and obtaining the purified recombined human tissue type plasminogen exciter TNK mutant rhTNK-tPA after elution by eluent c. The method has the characteristics of high capacity, high flow speed, easy regeneration, simple process and low cost, and is suitable for linear amplification and mass production.
Description
Technical field
The invention belongs to medical biotechnology recombinant protein separating and purifying technology field, be specifically related to the purification process of a kind of recombinant human histiotype plasminogen activator TNK mutant rhTNK-tPA.
Background technology
Recombinant human histiotype plasminogen activator TNK mutant (recombinant human tissue-type plasminogen activator for TNK mutant, rhTNK-tPA) be to utilize gene recombination technology in mammalian cell, to express the novel thrombolytic agent of producing, the clinical thromboembolism treatment that is used for Acute Myocardial Infarction is a thrombus dissolving first aid medicine the most safely and effectively so far.In the production process of rhTNK-tPA, purifying process has material impact to the quality of product.
The purification scheme of existing t-PA is primarily aimed at its molecular structure characteristics and determines.Utilize the iso-electric point pI of t-PA to adopt ion exchange chromatography such as SP-Sepharose, CM-Sepharose etc. than higher characteristics; Utilize Histidine in the t-PA molecule, halfcystine, the higher characteristics of tryptophane equal size can adopt metal chelate chromatography method such as Zn
2+-Sepharose, Zn
2+-POROS 20MC etc.; Because the t-PA molecule has a lysine-binding site can adopt Methionin affinity chromatography such as Lysine-Sepharose; Can adopt heparin affinity chromatography method such as Heparin-Sepharose at the heparin binding site, Heparin HyperD etc.Problems such as it is not high that aforesaid method all has carrying capacity, and flow velocity is unhappy, and regenerative process is loaded down with trivial details.Other method, can be used for purifying t-PA as monoclonal antibody affinity chromatography, benzenyl amidine affinity chromatography, hydrophobic affinity chromatography such as Phenyl-Sepharose etc., reversed phase chromatography method, sieve method such as Sephacryl S-200 etc., but consider t-PA preparation large usage quantity, monoclonal antibody affinity chromatography and sieve method not too are applicable to the large scale purification of t-PA.Benzenyl amidine is used for the purifying serine protease, and also the someone is used for purifying t-PA or separate strand, double-stranded t-PA, but because it can not clean with alkali, can not remove the processing of pyrogen, costs an arm and a leg in addition, and it is not ideal enough to be used for purifying t-PA.
Summary of the invention
The object of the present invention is to provide the purification process of a kind of recombinant human histiotype plasminogen activator TNK mutant rhTNK-tPA, this method carrying capacity height, flow velocity is fast, and regeneration is easy, and process is easy, cost is low.
Above-mentioned purpose of the present invention is achieved by the following technical solution: the purification process of a kind of recombinant human histiotype plasminogen activator TNK mutant rhTNK-tPA comprises following steps:
(1) gets supernatant liquor after the cell culture fluid filtration with rhTNK-tPA, behind Blue Sepharose 6FastFlow affinity column chromatography, with the elutriant a1 that must contain target peak behind the elutriant a wash-out;
(2) the elutriant a1 that contains target peak of step (1) gained is carried out Lysine HyperD affinity column chromatography after, must contain the elutriant b1 of target peak with elutriant b wash-out;
(3) the elutriant b1 that contains target peak that gets step (2) gained carries out Sephadex G-25 gel-filtration, the recombinant human histiotype plasminogen activator TNK mutant rhTNK-tPA after promptly getting purifying behind the elutriant c wash-out.
In the step of the present invention (1) when Blue Sepharose 6Fast Flow affinity column chromatography, by BlueSepharose 6 Fast Flow to the binding capacity of rhTNK-tPA in the 6-10mg/ml filler, the composition of elutriant a is: the sodium chloride solution of 20-100mM phosphate buffered saline buffer, 1-2M, 2-5M urea, 0.04% tween 80, the pH value of elutriant is 7.0-9.0, and the consumption of elutriant a is a 1.5-3 times of column volume.
In the step of the present invention (1) before carrying out Blue Sepharose 6Fast Flow affinity column chromatography, need Blue Sepharose 6Fast Flow chromatography column to be carried out balance and removal of impurities processing with damping fluid, the component of damping fluid is the tween 80 damping fluid of 10-100mM phosphoric acid salt and 0.04% during balance, its consumption is 1.5-3 a times of column volume, the component of damping fluid is 10-100mM phosphate buffered saline buffer, 0.04% tween 80 and the sodium chloride solution of 1-3M during removal of impurities, its consumption be column volume 1.5-3 doubly.
After need diluting 2-9 times, the elutriant a1 that the middle gained of step of the present invention (1) contains target peak carries out the LysineHyperD affinity column chromatography again.
In the step of the present invention (2) when carrying out Lysine HyperD affinity column chromatography, by Lysine HyperD to the binding capacity of rhTNK-tPA in the 6-10mg/ml filler, the composition of elutriant b is: 20-100mM phosphate buffered saline buffer, the amino n-caproic acid of 0.1-0.5M 6-, 0.1-0.5M arginine, 0.04% tween 80, the pH value of elutriant is 7.0-9.0, and the consumption of elutriant b is a 1.5-3 times of column volume.
In the step of the present invention (2) before carrying out Lysine HyperD affinity column chromatography, need the LysineHyperD chromatography column to be carried out balance and removal of impurities processing with damping fluid, the component of damping fluid is the tween 80 damping fluid of 10-100mM phosphoric acid salt and 0.04% during balance, its consumption is 1.5-3 a times of column volume, the component of the damping fluid during removal of impurities is 10-100mM phosphate buffered saline buffer, 0.04% tween 80 and the sodium chloride solution of 1-3M, its consumption be column volume 1.5-3 doubly.
When carrying out Sephadex G-25 gel-filtration, the component of elutriant c is 0.1-0.5M arginine and 0.04%Tween 80 in the step of the present invention (3), and the pH value is 7.0-9.0, and the consumption of elutriant c is a 1.5-3 times of column volume.
In the step of the present invention (3) before carrying out Sephadex G-25 gel-filtration, need the SephadexG-25 chromatography column to be carried out Balance Treatment with damping fluid, the component of the damping fluid during balance is 0.1-0.5M arginine and 0.04% tween 80, the pH value is 7.0-9.0, and the consumption of damping fluid is a 1.5-3 times of column volume.
The purity of recombinant human histiotype plasminogen activator TNK mutant rhTNK-tPA behind the purifying of step of the present invention (3) preparation is greater than 98%.
Purifying principle of the present invention is: rhTNK-tPA is a glycosylated protein, can combine with Blue Sepharose 6FastFlow filler specificity; RhTNK-tPA contains the Methionin pocket, can combine with lysine HyperD specificity; Its molecular weight is 68KDa, can filter by Sephadex G-25Medium and carry out desalination.Be specially: be primarily aimed at its molecular structure characteristics according to the purification scheme of rhTNK-tPA and determine, overall plan is to concentrate thick purifying earlier fast, carry out consummateization again.RhTNK-tPA is by expressing cho cell, and the relative concentration of target protein in cells and supernatant is lower, adds that sample volume is big, and the first step purification process is must treatment capacity big, and speed is fast.Reach this purpose, the chromatographic stuffing of use must possess the big and fast characteristics of flow velocity of binding capacity to rhTNK-tPA.The carrying capacity of Blue Sepharose 6Fast Flow filler is 6-10mg/ml, and its wash water, equilibrium velocity are 200cm/h, and last sample flow velocity is 180cm/h; Compare with fillers such as SP-Sepharose, Zn2+-Sepharose, Zn2+-POROS 20M, it is higher that it has carrying capacity, faster, the characteristics more easily of regenerating of flow velocity.Consummateization in second step must have extraordinary specificity to target protein, at the Methionin pocket that contains among the t-PA, generally all can use the Methionin filler of being correlated with, by contrast Lysine-Sepharose and Lysine HyperD, find that the carrying capacity of Lysine HyperD is higher, flow velocity is faster; During consummateization of this external application Lysine HyperD, the purity of rhTNK-tPA is higher.Because the elution peak damping fluid of Lysine HyperD chromatography does not meet the requirement of these goods product formula, so utilize Sephadex G-25 to carry out desalting treatment again, rhTNK-tPA is replaced final damping fluid.
Compared with prior art, the present invention has following advantage:
Characteristics such as (1) Blue Sepharose 6Fast Flow chromatography has the carrying capacity height, and flow velocity is fast, and regeneration is convenient, and concentrated and purification effect is remarkable, through Blue Sepharose 6Fast Flow chromatography, sample volume can dwindle more than 100 times;
(2) the purification efficiency height of rhTNK-tPA, the rhTNK-tPA behind the purifying accounts for more than 98% of total protein concentration, and monomer purity reaches 500,000 IU/mg greater than 95% than living;
(3) simple to operate, good reproducibility, but linear amplification, damping fluid is raw materials used cheap, can be used for scale operation.
Embodiment
The present invention is further illustrated below in conjunction with specific embodiment, and following reagent is commercially available if no special instructions.
Embodiment 1
The purification process of the recombinant human histiotype plasminogen activator TNK mutant rhTNK-tPA that present embodiment provides is as follows:
(1) supernatant pre-treatment: the serum-free culture supernatant of harvested cell in reactor, remove cell debris with filtering with microporous membrane, with protection purifying filler;
(2) Blue Sepharose 6Fast Flow affinity column chromatography: with 2.5 times of column volumes of buffer A balance, the component of buffer A is 20mM phosphate buffered saline buffer and 0.04% tween 80, pH7.2; Last sample; With 2.5 times of column volumes of buffer A flushing; Wash out foreign protein with buffer B, the component of buffer B is to add 2M NaCl on the basis of buffer A; Use the damping fluid C wash-out of 2.5 times of column volumes to obtain target peak, the component of damping fluid C is 20mM phosphoric acid salt, 1MNaCl, 2M urea and 0.04% tween 80, pH7.20; Target peak carries out Lysine HyperD chromatography after using phosphate buffered saline buffer to dilute 2 times;
(3) Lysine HyperD affinity column chromatography: balance is consistent with Blue Sepharose 6Fast Flow chromatography with last sample process, use the damping fluid D of 2.5 times of column volumes to wash out foreign protein after finishing, the component of damping fluid D is to add 1M NaCl on the basis of buffer A; Use the damping fluid E wash-out of 2.5 times of column volumes to obtain target peak, the component of damping fluid E is the amino n-caproic acid of 20mM phosphoric acid salt, 0.1M 6-, 0.2M arginine, 0.04% tween 80, pH7.0; Target peak carries out Sephadex G-25Medium gel-filtration;
(4) Sephadex G-25Medium gel-filtration: use 2.5 times of column volume damping fluid F to carry out balance, the component of damping fluid F is 0.1M arginine and 0.04% tween 80, pH7.0; Use the sample of 25% column volume to go up sample, with damping fluid F target protein is eluted, obtain satisfactory rhTNK-tPA solution, the purity of the recombinant human histiotype plasminogen activator TNK mutant behind the purifying of preparation is greater than 98%.
In the damping fluid of all uses, all contain volumn concentration and be 0.04% tween 80.
Embodiment 2
The purification process of the recombinant human histiotype plasminogen activator TNK mutant rhTNK-tPA that present embodiment provides is as follows:
(1) gets supernatant liquor after the cell culture fluid filtration with rhTNK-tPA, behind Blue Sepharose 6FastFlow affinity column chromatography, with the elutriant a1 that must contain target peak behind the elutriant a wash-out;
(2) the elutriant a1 that contains target peak of step (1) gained is carried out Lysine HyperD affinity column chromatography after, must contain the elutriant b1 of target peak with elutriant b wash-out;
(3) the elutriant b1 that contains target peak that gets step (2) gained carries out Sephadex G-25 gel-filtration, the recombinant human histiotype plasminogen activator TNK mutant rhTNK-tPA after promptly getting purifying behind the elutriant c wash-out.
In above-mentioned steps:
In the step (1) when Blue Sepharose 6Fast Flow affinity column chromatography, by Blue Sepharose 6Fast Flow to the binding capacity of rhTNK-tPA in the 6-10mg/ml filler, the composition of elutriant a is: the sodium chloride solution of 50mM phosphate buffered saline buffer, 1.5M, 3M urea, 0.04% tween 80, the pH value of elutriant is 8.0, and the consumption of elutriant a is 2 times of column volumes.
In the step (1) before carrying out Blue Sepharose 6Fast Flow affinity column chromatography, need Blue Sepharose 6Fast Flow chromatography column to be carried out balance and removal of impurities processing with damping fluid, the component of damping fluid is the tween 80 damping fluid of 50mM phosphoric acid salt and 0.04% during balance, its consumption is 2 times of column volume, the component of damping fluid is 50mM phosphate buffered saline buffer, 0.04% tween 80 and the sodium chloride solution of 1.5M during removal of impurities, and its consumption is 2 times of column volume.
After need diluting 5 times, the elutriant a1 that the middle gained of step (1) contains target peak carries out Lysine HyperD affinity column chromatography again.
In the step (2) when carrying out Lysine HyperD affinity column chromatography, by Lysine HyperD to the binding capacity of rhTNK-tPA in the 6-10mg/ml filler, the composition of elutriant b is: 50mM phosphate buffered saline buffer, the amino n-caproic acid of 0.3M 6-, 0.3M arginine, 0.04% tween 80, the pH value of elutriant is 8.0, and the consumption of elutriant b is 2 times of column volumes.
In the step (2) before carrying out Lysine HyperD affinity column chromatography, need Lysine HyperD chromatography column to be carried out balance and removal of impurities processing with damping fluid, the component of damping fluid is the tween 80 damping fluid of 50mM phosphoric acid salt and 0.04% during balance, its consumption is 2 times of column volume, the component of the damping fluid during removal of impurities is 50mM phosphate buffered saline buffer, 0.04% tween 80 and the sodium chloride solution of 1.5M, and its consumption is 2 times of column volume.
When carrying out Sephadex G-25 gel-filtration, the component of elutriant c is 0.3M arginine and 0.04%Tween 80 in the step (3), and the pH value is 8.0, and the consumption of elutriant c is 2 times of column volumes.
In the step (3) before carrying out Sephadex G-25 gel-filtration, need Sephadex G-25 chromatography column to be carried out Balance Treatment with damping fluid, the component of the damping fluid during balance is 0.3M arginine and 0.04% tween 80, and the pH value is 8.0, and the consumption of damping fluid is 2 times of column volumes.
The purity of recombinant human histiotype plasminogen activator TNK mutant rhTNK-tPA behind the purifying of step (3) preparation is greater than 98%.
Embodiment 3
The purification process of the recombinant human histiotype plasminogen activator TNK mutant rhTNK-tPA that present embodiment provides is as follows:
(1) supernatant pre-treatment: the serum-free culture supernatant of harvested cell in reactor, remove cell debris with filtering with microporous membrane, with protection purifying filler;
(2) get the rhTNK-tPA supernatant liquor, behind Blue Sepharose 6Fast Flow affinity column chromatography, with the elutriant a1 that must contain target peak behind the elutriant a wash-out;
(3) after the elutriant a1 that step (1) gained is contained target peak carries out Lysine HyperD affinity column chromatography, must contain the elutriant b1 of target peak with elutriant b wash-out;
(4) get the elutriant b1 that step (2) gained contains target peak and carry out Sephadex G-25 gel-filtration, the recombinant human histiotype plasminogen activator TNK mutant after promptly getting purifying behind the elutriant c wash-out.
In above-mentioned steps:
In the step (2) when Blue Sepharose 6Fast Flow affinity column chromatography, by Blue Sepharose 6FastFlow to the binding capacity of rhTNK-tPA in the 6-10mg/ml filler, the composition of elutriant a is: the sodium chloride solution of 100mM phosphate buffered saline buffer, 2M, 5M urea, 0.04% tween 80, the pH value of elutriant is 9.0, and the consumption of elutriant a is 1.5 times of column volumes.
In the step (2) before carrying out Blue Sepharose 6Fast Flow affinity column chromatography, need Blue Sepharose 6Fast Flow chromatography column to be carried out balance and removal of impurities processing with damping fluid, the component of damping fluid is the tween 80 damping fluid of 100mM phosphoric acid salt and 0.04% during balance, its consumption is 1.5 times of column volume, the component of damping fluid is 100mM phosphate buffered saline buffer, 0.04% tween 80 and the sodium chloride solution of 3M during removal of impurities, and its consumption is 1.5 times of column volume.
After need diluting 9 times, the elutriant a1 that the middle gained of step (2) contains target peak carries out Lysine HyperD affinity column chromatography again.
In the step (3) when carrying out Lysine HyperD affinity column chromatography, by Lysine HyperD to the binding capacity of rhTNK-tPA in the 6-10mg/ml filler, the composition of elutriant b is: 100mM phosphate buffered saline buffer, the amino n-caproic acid of 0.5M 6-, 0.5M arginine, 0.04% tween 80, the pH value of elutriant is 9.0, and the consumption of elutriant b is 1.5 times of column volumes.
In the step (3) before carrying out Lysine HyperD affinity column chromatography, need Lysine HyperD chromatography column to be carried out balance and removal of impurities processing with damping fluid, the component of damping fluid is the tween 80 damping fluid of 100mM phosphoric acid salt and 0.04% during balance, its consumption is 1.5 times of column volume, the component of the damping fluid during removal of impurities is 100mM phosphate buffered saline buffer, 0.04% tween 80 and the sodium chloride solution of 3M, and its consumption is 1.5 times of column volume.
When carrying out Sephadex G-25 gel-filtration, the component of elutriant c is 0.5M arginine and 0.04%Tween 80 in the step (4), and the pH value is 8.5, and the consumption of elutriant c is 1.5 times of column volumes.
In the step (4) before carrying out Sephadex G-25 gel-filtration, need Sephadex G-25 chromatography column to be carried out Balance Treatment with damping fluid, the component of the damping fluid during balance is 0.5M arginine and 0.04% tween 80, and the pH value is 9.0, and the consumption of damping fluid is 1.5 times of column volumes.
The purity of recombinant human histiotype plasminogen activator TNK mutant rhTNK-tPA behind the purifying of step (4) preparation is greater than 98%.
The foregoing description is a preferred implementation of the present invention; but embodiments of the present invention are not restricted to the described embodiments; other any do not deviate from change, the modification done under spirit of the present invention and the principle, substitutes, combination, simplify; all should be the substitute mode of equivalence, be included in protection scope of the present invention.
Claims (9)
1. the purification process of a recombinant human histiotype plasminogen activator TNK mutant rhTNK-tPA is characterized in that comprising following steps:
(1) gets supernatant liquor after the cell culture fluid filtration with rhTNK-tPA, behind Blue Sepharose 6FastFlow affinity column chromatography, with the elutriant a1 that must contain target peak behind the elutriant a wash-out;
(2) the elutriant a1 that contains target peak of step (1) gained is carried out Lysine HyperD affinity column chromatography after, must contain the elutriant b1 of target peak with elutriant b wash-out;
(3) the elutriant b1 that contains target peak that gets step (2) gained carries out Sephadex G-25 gel-filtration, the recombinant human histiotype plasminogen activator TNK mutant rhTNK-tPA after promptly getting purifying behind the elutriant c wash-out.
2. purification process according to claim 1, it is characterized in that: in the step (1) when Blue Sepharose6Fast Flow affinity column chromatography, by Blue Sepharose 6Fast Flow to the binding capacity of rhTNK-tPA in the 6-10mg/ml filler, the composition of elutriant a is: the sodium chloride solution of 20-100mM phosphate buffered saline buffer, 1-2M, 2-5M urea, 0.04% tween 80, the pH value of elutriant is 7.0-9.0, and the consumption of elutriant a is a 1.5-3 times of column volume.
3. purification process according to claim 2, it is characterized in that: in the step (1) before carrying out Blue Sepharose 6Fast Flow affinity column chromatography, need Blue Sepharose 6Fast Flow chromatography column to be carried out balance and removal of impurities processing with damping fluid, the component of damping fluid is the tween 80 of 10-100mM phosphate buffered saline buffer and 0.04% during balance, its consumption is 1.5-3 a times of column volume, the component of damping fluid is 10-100mM phosphate buffered saline buffer, 0.04% tween 80 and the sodium chloride solution of 1-3M during removal of impurities, its consumption be column volume 1.5-3 doubly.
4. purification process according to claim 1 is characterized in that: carry out Lysine HyperD affinity column chromatography again after the elutriant a1 that the middle gained of step (1) contains target peak need dilute 2-9 times.
5. purification process according to claim 1, it is characterized in that: in the step (2) when carrying out Lysine HyperD affinity column chromatography, by Lysine HyperD to the binding capacity of rhTNK-tPA in the 6-10mg/ml filler, the composition of elutriant b is: 20-100mM phosphate buffered saline buffer, the amino n-caproic acid of 0.1-0.5M 6-, 0.1-0.5M arginine, 0.04% tween 80, the pH value of elutriant is 7.0-9.0, and the consumption of elutriant b is a 1.5-3 times of column volume.
6. purification process according to claim 5, it is characterized in that: in the step (2) before carrying out the LysineHyperD affinity column chromatography, need Lysine HyperD chromatography column to be carried out balance and removal of impurities processing with damping fluid, the component of damping fluid is the tween 80 of 10-100mM phosphate buffered saline buffer and 0.04% during balance, its consumption is 1.5-3 a times of column volume, the component of the damping fluid during removal of impurities is 10-100mM phosphate buffered saline buffer, 0.04% tween 80 and the sodium chloride solution of 1-3M, its consumption be column volume 1.5-3 doubly.
7. purification process according to claim 1, it is characterized in that: in the step (3) when carrying out the SephadexG-25 gel-filtration, the component of elutriant c is 0.1-0.5M arginine and 0.04%Tween 80, and the pH value is 7.0-9.0, and the consumption of elutriant c is a 1.5-3 times of column volume.
8. purification process according to claim 7, it is characterized in that: in the step (3) before carrying out the SephadexG-25 gel-filtration, need Sephadex G-25 chromatography column to be carried out Balance Treatment with damping fluid, the component of the damping fluid during balance is 0.1-0.5M arginine and 0.04% tween 80, the pH value is 7.0-9.0, and the consumption of damping fluid is a 1.5-3 times of column volume.
9. purification process according to claim 1 is characterized in that: the purity of the recombinant human histiotype plasminogen activator TNK mutant rhTNK-tPA behind the purifying of step (3) preparation is greater than 98%.
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| CN102199587A (en) * | 2011-03-24 | 2011-09-28 | 广东药学院 | Functional mutant of human plasminogen, its preparation method and application |
| CN108424897A (en) * | 2018-03-13 | 2018-08-21 | 广州铭康生物工程有限公司 | A kind of purification process of rhTNK-tPA cells harvest liquid |
| CN108704122A (en) * | 2018-05-23 | 2018-10-26 | 广州铭康生物工程有限公司 | A kind of injection rhTNK-tPA lyophilized preparations and preparation method thereof |
| CN113025604A (en) * | 2021-03-23 | 2021-06-25 | 重庆中元汇吉生物技术有限公司 | Reagents and methods for preparing high purity PICs |
| CN113337490A (en) * | 2021-06-18 | 2021-09-03 | 广州铭康生物工程有限公司 | Method for large-scale rapid separation and purification of rhTNK-tPAI/II type |
| CN114426962A (en) * | 2021-12-21 | 2022-05-03 | 佛山汉腾生物科技有限公司 | t-PA purification process |
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| CN102199587A (en) * | 2011-03-24 | 2011-09-28 | 广东药学院 | Functional mutant of human plasminogen, its preparation method and application |
| CN102199587B (en) * | 2011-03-24 | 2013-06-19 | 广东药学院 | Functional mutant of human plasminogen, its preparation method and application |
| CN108424897A (en) * | 2018-03-13 | 2018-08-21 | 广州铭康生物工程有限公司 | A kind of purification process of rhTNK-tPA cells harvest liquid |
| CN108424897B (en) * | 2018-03-13 | 2020-08-14 | 广州铭康生物工程有限公司 | Purification method of rhTNK-tPA cell harvest liquid |
| CN108704122A (en) * | 2018-05-23 | 2018-10-26 | 广州铭康生物工程有限公司 | A kind of injection rhTNK-tPA lyophilized preparations and preparation method thereof |
| CN113025604A (en) * | 2021-03-23 | 2021-06-25 | 重庆中元汇吉生物技术有限公司 | Reagents and methods for preparing high purity PICs |
| CN113337490A (en) * | 2021-06-18 | 2021-09-03 | 广州铭康生物工程有限公司 | Method for large-scale rapid separation and purification of rhTNK-tPAI/II type |
| CN114426962A (en) * | 2021-12-21 | 2022-05-03 | 佛山汉腾生物科技有限公司 | t-PA purification process |
| CN114426962B (en) * | 2021-12-21 | 2024-06-18 | 佛山汉腾生物科技有限公司 | T-PA purification method |
| CN115475258A (en) * | 2022-09-06 | 2022-12-16 | 江苏丰华生物制药有限公司 | Purification process of tenecteplase for injection |
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