A kind of cleaning method of affinity chromatography column
A chromatographic column and affinity technology, applied in chemical instruments and methods, separation methods, solid adsorbent liquid separation, etc., can solve problems such as inability to guarantee the cleaning effect
Active Publication Date: 2018-12-04
TASLY BIOPHARMACEUTICALS CO LTD
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Problems solved by technology
[0003] The existing cleaning methods are mainly soaking and rinsing with water, or soaking and rinsing with buffer solution, acid, alkali and other solutions, but because there is no proper detection method to detect the cleaned packing, the cleaning effect cannot be guaranteed
Method used
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Embodiment 1
[0162] Wash the used affinity chromatography column (filler is Benzamidine Sepharose4FF H-SUB, column model: BPG100, height 15cm, diameter 10cm) with 4200ml of water for injection, then with 7000ml of 6M guanidine hydrochloride, then with 4200ml of water for injection , wash with 4200ml 0.5M NaOH, then wash the column with 14000ml0.01mol / L equilibrium buffer (pH7.0±0.1 containing 0.4M NaCl), the liquid flow rate during cleaning is 120ml / min, take 10ml of cleaning solution in a test tube, Drop 2-3 drops of phosphoric acid and observe whether there is precipitation. If there is precipitation, it must be diluted with MilliQ water until no precipitation appears after adding phosphoric acid. Three times, if the results of the three measurements are all lower than 500ppm, it means that the cleaning is qualified; after the cleaning is qualified, the affinity chromatography column is stored in a chromatographic cabinet at 2-12°C.
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The invention relates to a washing method of affinity chromatography column. The washing method comprises the following steps: step one, filling materials into a dirty affinity chromatography column, washing the affinity chromatography column by water, and then washing the affinity chromatography column by guanidine hydrochloride; step two, washing the affinity chromatography column by water, and then washing the affinity chromatography column by a water solution of sodium hydroxide; step three, washing the affinity chromatography column by an equilibrium buffer solution with a pH value of 5.0-8.0; and according to the actual needs, the washing method may also comprise step four and step five: step four, dropwise adding phosphoric acid into the washing liquid obtained in the step three, observing whether precipitate is generated or not, if no precipitate is generated, measuring the washing liquid by a total organic carbon analyzer, wherein if the measurement result is less than 500 ppm, the washing is qualified; step five, if precipitate is generated, diluting the washing liquid obtained in the step four by water until no precipitate is generated after the phosphoric acid is dropwise added, and measuring the diluted washing liquid by a total organic carbon analyzer, wherein if the data obtained by multiplying the measurement result by the dilution times is less than 500 ppm, the washing is qualified.
Description
technical field [0001] The invention relates to a cleaning method for a chromatographic column, in particular to a cleaning method for an affinity chromatographic column. Background technique [0002] Affinity chromatography is a technique for protein separation, which can adsorb proteins with special structures on chromatographic columns to achieve the purpose of protein purification. In the field of biopharmaceuticals, affinity chromatography is widely used. After the affinity chromatography column is used, it may pollute the next use and introduce foreign pollutants, such as introducing endotoxin to the next use or introducing proteins that were not cleaned last time into the next product. There is no fixed technical scheme for the cleaning method of the existing affinity chromatography column, so the cleaning effect cannot be guaranteed. In order to ensure that the affinity chromatography column is cleaned after each use and will not pollute the next use, it is necessa...
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IPC IPC(8): B01D15/38B01D15/42
Inventor 李雪峰赵国焓邵海玲
Owner TASLY BIOPHARMACEUTICALS CO LTD