Method for detecting purity of protein sample containing polyethylene glycol by using capillary electrophoresis technology

A capillary electrophoresis, polyethylene glycol technology, applied in the preparation of test samples, material inspection products, measuring devices, etc., can solve the problem of little optimization experience and technology, no suitable capillary electrophoresis analysis method, affecting the purity and accuracy of protein samples. Analysis and other problems to achieve the effect of preventing fragmentation and insufficient denaturation

Active Publication Date: 2020-06-02
SHANGHAI WUXI BIOLOGIC TECH CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

When using a capillary electrophoresis instrument to detect a protein sample containing a certain concentration of polyethylene glycol, there will be aggregate peaks that cannot be confirmed by other technical means in the test result spectrum, which will affect the accurate analysis of the purity of the protein sample
At present, there are no reports and solutions for the impact of polyethylene glycol on capillary electrophoresis detection, and the purpose of the present invention is to solve this problem
[0004] For the booming biopharmaceutical industry, the emergence of different process development systems will inevitably bring more factors affecting the purity analysis, but there are no reports on capillary electrophoresis analysis methods suitable for different process development systems. Most of the biopharmaceuticals in China Manufacturers rarely have the experience and technology to independently optimize this method, so related work is necessary

Method used

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  • Method for detecting purity of protein sample containing polyethylene glycol by using capillary electrophoresis technology
  • Method for detecting purity of protein sample containing polyethylene glycol by using capillary electrophoresis technology
  • Method for detecting purity of protein sample containing polyethylene glycol by using capillary electrophoresis technology

Examples

Experimental program
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Effect test

Embodiment 1

[0048] Preparation of buffer solution: Take disodium hydrogen phosphate and citric acid solid powder, add pure water to prepare a buffer solution containing 15.8mmol / L disodium hydrogen phosphate and 4.9mmol / L citric acid.

[0049] The method of the present invention: the protein sample to be analyzed is Herceptinumab solution (21 mg / mL), and the final concentration of polyethylene glycol 8000 is set to 2% (P=2). Substitute the P value and the HMW value (HMW=0.15) of Herceptinumab solution (21 mg / mL) without PEG into HMW=-110.1+1.576T+21.1U+139.12P-0.296T×U In the model of -2.058T×P-30.75U×P+0.4502T×U×P, the value of T should be as low as possible, and the final concentration of urea required was calculated using Minitab software: 3mol / L, and the incubation temperature was: 64.5°C.

[0050] Add the corresponding mass of polyethylene glycol 8000, urea, and sodium lauryl sulfate to the protein sample, and use buffer solution to adjust the total volume of the protein sample to 10...

Embodiment 2

[0056] Preparation of buffer solution: Take disodium hydrogen phosphate and citric acid solid powder, add pure water to prepare a buffer solution containing 15.8mmol / L disodium hydrogen phosphate and 4.9mmol / L citric acid.

[0057] The method of the present invention: the protein to be analyzed is Herceptinumab solution (21 mg / mL), and the final concentration of polyethylene glycol 8000 is set to 4% (P=4). Substitute the P value and the HMW value (HMW=0.15) of Herceptinumab solution (21 mg / mL) without PEG into HMW=-110.1+1.576T+21.1U+139.12P–0.296T×U In the model of –2.058T×P–30.75U×P+0.4502T×U×P, T should be as low as possible, and the final concentration of urea required was calculated using Minitab software: 3mol / L, and the incubation temperature was: 65.0°C.

[0058] Add the corresponding mass of polyethylene glycol 8000, urea and sodium lauryl sulfate to the sample and use a buffer solution to adjust the total sample volume to 100 μL and adjust the concentration of Hercep...

Embodiment 3

[0064] Preparation of buffer solution: Take disodium hydrogen phosphate and citric acid solid powder, add pure water to prepare a buffer solution containing 15.8mmol / L disodium hydrogen phosphate and 4.9mmol / L citric acid.

[0065] The method of the present invention: the protein to be analyzed is Herceptinumab solution (21 mg / mL), and the final concentration of polyethylene glycol 8000 is set to 6% (P=6). Substitute the P value and the HMW value (HMW=0.15) of Herceptinumab solution (21 mg / mL) without PEG into HMW=2954–94.3T–89.1U+112.1P+0.7489T×T+ In the 1.304T×U–1.659T×P model, T should be as low as possible, and the required final concentration of urea was calculated using Minitab software: 3mol / L, and the incubation temperature was: 65.6°C.

[0066] Add the corresponding mass of polyethylene glycol 8000, urea and sodium lauryl sulfate to the sample and use a buffer solution to adjust the total sample volume to 100 μL and adjust the concentration of Herceptin to 1 mg / mL, po...

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Abstract

The invention discloses a method for detecting the purity of a protein sample containing polyethylene glycol by using a capillary electrophoresis technology. The method comprises the following steps of (1) adding urea and related reagents into the protein sample for pretreatment; (2) incubating the sample at a proper temperature; (3) carrying out capillary electrophoresis analysis; and (4) calculating the purity of the protein sample according to the peak area ratio of different components, wherein the final concentration of urea and the incubation temperature are determined by a model. By adding urea and optimizing the incubation temperature, the problems of peak shape deformation, difficulty in accurate quantification and the like when the purity of a drug is measured by the traditionalcapillary electrophoresis are solved, and the accuracy and precision of the purity measurement are guaranteed.

Description

technical field [0001] The invention relates to the field of biological separation analysis, in particular to a method for detecting the purity of a protein sample containing polyethylene glycol by using capillary electrophoresis technology. Background technique [0002] In the production and development of protein samples, different process systems will produce more process-related additives, which come from various links in the production process, such as in order to improve the resolution and maintenance of protein fragments with different molecular weights in the downstream purification process and drug formulation development process. Protein molecules are stabilized by adding polyethylene glycol. However, the introduction of these additives may affect the detection of protein sample purity during sample pretreatment, instrument injection analysis and other stages, so that true and accurate purity detection results cannot be obtained. Therefore, in order to ensure that...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N27/447G01N33/68G01N1/38
CPCG01N27/447G01N33/6803G01N1/38
Inventor 麻旭田橙屈晨张亚楠丁晓娜沈璇武涛时昕李志国曹晓林黄岗周伟昌陈智胜
Owner SHANGHAI WUXI BIOLOGIC TECH CO LTD
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