CN104922936B - A kind of cleaning method of cation-exchange chromatography post - Google Patents
A kind of cleaning method of cation-exchange chromatography post Download PDFInfo
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Abstract
The present invention relates to a kind of cleaning methods of cation-exchange chromatography post, and described method includes following steps：（1）It by used cation-exchange chromatography post, washes with water, then uses NaOH solution（2）Pillar finally is rushed with equilibration buffer (pH5.0-8), receives cleaning solution.The invention also includes steps（3）With（4）Verification step, it is specific as follows：（3）Cleaning solution is taken, phosphoric acid is dripped, whether there is or not precipitatings to generate for observation, such as without precipitating, measures cleaning solution with total organic carbon analyzer, measurement result is lower than 500ppm, then it represents that cleaning is qualified.（4）If there is precipitating to generate, after occurring after step 2 must being obtained cleaning solution dilution until phosphoric acid is added dropwise with water without precipitating, cleaning solution after dilution is measured into the cleaning solution with total organic carbon analyzer, the data that measurement result is obtained multiplied by extension rate are lower than 500ppm, then it represents that cleaning is qualified.
Description
Technical field
The present invention relates to the cleaning method of chromatographic column, in particular to a kind of cleaning method of cation-exchange chromatography post.
Background technique
In field of biological pharmacy, cation-exchange chromatography post is widely used.After cation-exchange chromatography post use, fill out
Some residuals are had in material, next use may be polluted, introducing external contaminant, such as can be to making next time
With being introduced into endotoxin or the albumen that does not clean up of last time be introduced into next product.
Existing cleaning method is mainly soaked in water flushing, or with buffer solution, acid, and the solution such as alkali carry out soaking flushing, but
Because being detected without detection method appropriate to the filler cleaned, cleaning effect can not be ensured.To guarantee to use every time
Cation-exchange chromatography post afterwards cleans up, and will not hand over the cation for using polluting, needing to establish specification next time
Change the cleaning process of chromatographic column.
The present invention after study, finds a kind of simple and easy, is suitble to industrialized cleaning method, i.e., is cleaned by three steps
Method, it is ensured that endotoxin and total organic carbon cleaning are qualified.The present invention hands over cation by cleaning method that is fixed and optimizing
It changes chromatographic column to be cleaned, so that cation-exchange chromatography post cleaning effect is ensured, eliminates cation-exchange chromatography
Interior endotoxin, microorganism and last time use remaining albumen, it is ensured that total organic carbon is examined qualified.
Summary of the invention
The present invention provides a kind of cleaning method of cation-exchange chromatography post, the cleaning refers to using or will
The unclean filler used is cleaned, and column chromatography next time is carried out after cleaning is qualified.
Cation-exchange chromatography post of the present invention, filler are any filling out with cation-exchange chromatography function
Material, preferably：SP Sepharose Fast Flow, S Sepharose Fast Flow is most preferably SP Sepharose
Fast Flow。
Above-mentioned filler can be by prior art preparation, can also be by being commercially available.
The present invention provides a kind of cleaning method of cation-exchange chromatography post, includes the following steps：
Step 1,
It by unclean cation-exchange chromatography post filler, washes with water, then with NaOH aqueous cleaning；
Step 2,
It is cleaned with equilibration buffer (pH5.0-8).
As needed, the step of the invention also includes step 3 and steps 4, it is as follows：
Step 3,
The cleaning solution that step 2 obtains is taken, phosphoric acid is dripped, whether there is or not precipitatings to generate for observation, such as without precipitating, uses total organic carbon analyzer
The cleaning solution is measured, measurement result is lower than 500ppm, then it represents that cleaning is qualified.
Step 4,
If there is precipitating to generate, after must occurring cleaning solution dilution without precipitating after phosphoric acid is added dropwise with water, after dilution
Cleaning solution measures the cleaning solution with total organic carbon analyzer, and measurement result is lower than 500ppm multiplied by the data that extension rate obtains,
Then indicate that cleaning is qualified
Cation-exchange chromatography post of the present invention, preferred filler are SP Sepharose Fast Flow；Column type number：
It is a kind of in BPG100, BPG500, XK2, XK20, XK50, XK20, preferably BPG100.The high ratio of the pillar diameter：（0.5-1）：1,
It is preferred that（0.5-0.7）：1.Within the most preferably high 15cm of pillar, diameter：10cm.
Cleaning method of the invention cleans in a reservoir after can removing filler from chromatographic column, can also be mounted in chromatography
It is cleaned in column, is preferably mounted in chromatographic column and cleans, wherein cleaning solution flow velocity is 120-150ml/min, preferably 120ml/min.
Step of the present invention（1）Described in water be preferably water for injection, scavenging period 30-45min.
The sodium hydrate aqueous solution of the preferred 1-1.5M of the sodium hydrate aqueous solution, scavenging period 90min or more, most preferably
90-100min is cleaned for 1.0M sodium hydroxide.
The 1M sodium hydroxide preparation method：Sodium hydroxide 400.0g is weighed, moves it into serum bottle with water for injection
Or in liquid storing bag, it is put into stirrer, moving on magnetic stirring apparatus is uniformly dissolved solid all, will be molten in serum bottle or liquid storing bag
Liquid calibration continues stirring 2 minutes to 10L.Be placed in after preparation between the preparation of 2~12 DEG C of environment specify it is spare in region.
Step of the present invention（2）Described in equilibration buffer be preferably phosphate buffer and containing NaCl, preferably 0.01mol/L
Phosphate buffer（PH6.0 NaCl containing 0.1-0.2M）, most preferably 0.01mol/L phosphate buffer（PH6.0 NaCl containing 0.1M）.
It is preferred that the preparation of the 0.01mol/L equilibration buffer (pH6.0 ± 0.1 NaCl containing 0.1mol/L)：It weighs
Sodium chloride 58.9g, sodium dihydrogen phosphate 12.0g, disodium hydrogen phosphate 2.0g move it into serum bottle or liquid storing bag with water for injection
In, it is put into stirrer, moving on magnetic stirring apparatus is uniformly dissolved solid all, and standardization of solution in serum bottle or liquid storing bag is arrived
10L continues stirring 2 minutes, determines its pH value in 6.0 ± 0.1 ranges.It is placed in after preparation between the preparation of 2~12 DEG C of environment
It is spare in specified region.
Step of the present invention（3）Described in drop phosphoric acid, be that cleaning solution is added in container, the phosphoric acid of concentration be added, stand,
Visually observe that whether there is or not Precipitations.The purpose is to tentatively confirm phosphorus content in cleaning solution, if there is precipitating illustrate phosphorus content compared with
Height may can block total organic carbon analyzer pipeline in checkout procedure.
It is described to be measured with total organic carbon analyzer, it is with the organic carbon concentration in total organic carbon analyzer cleaning solution.This
Inventive step（4）Described in cleaning solution must be diluted with water, method is as follows：It, must be with water by cleaning solution dilution if there is precipitating to generate
Until the cleaning solution after dilution is measured the cleaning solution with total organic carbon analyzer after occurring after phosphoric acid is added dropwise without precipitating, measure
As a result it is lower than 500ppm multiplied by the data that extension rate obtains, then it represents that cleaning is qualified.
The explanation of relational language of the present invention：
1, cation-exchange chromatography：It is a kind of technology of Protein Separation, relies primarily on the interaction between charge, utilize band
Charge fine difference carries out Separation of Proteins in electric molecule.
2, endotoxin：It is one of gram-negative bacterial cell wall ingredient, is called lipopolysaccharides.Lipopolysaccharides is to host
It is virose.Endotoxin is only worked as bacterial death dissolution or is just released after destroying bacterium cell by artificial means.
3, total organic carbon（TOC）：Refer to the total amount that dissolubility and suspension organic matter are carbon containing in water body.TOC is one fast
The overall target of speed calibrating, it indicates that the total amount in water containing organic matter, unit are ppm or ppb with the quantity of carbon.In the world
TOC is by an important reference indicator as organic pollution degree in evaluation water body.
The cleaning method of cation-exchange chromatography post of the invention is obtained by screening, and screening process is as follows：
1, the selection of concentration of lotion
The following table 1 cleaning solution according to embodiment sequential irrigation, it is as a result as follows：
Table 1
3, the selection of cleaning solution time
Table 2
| Cleaning solution title | Scavenging period 1 | Scavenging period 2 | 3 scavenging period 4 of scavenging period |
| Sodium hydroxide | 60min | 90min | 100min110min |
| As a result | |||
| Endotoxin | > 0.25EU/ml | ≤0.25EU/ml | ≤0.25EU/ml≤0.25EU/ml |
| Total organic carbon | 2013ppb | 108ppb | 110ppb117ppb |
From upper table 2 as it can be seen that cleaning solution is identical with the effect of time 100 and 110, for saving principle, preferably cleaning solution time
For 90-100min.
Beneficial effects of the present invention are proved below by way of comparative experiments：
The prior art is rushed pillar method and is compared with method of the invention
Table 3
The prior art rushes pillar method 1:
Used cation-exchange chromatography post is cleaned into 30min with water for injection first, is then cleaned with 1M NaOH
60min finally rushes pillar 90min with 0.01mol/L equilibration buffer (pH6.0 ± 0.1 NaCl containing 0.1mol/L), when cleaning
Flow rate of liquid is：120ml/min；Pillar efflux is taken to carry out endotoxin and TOC detection, it is as a result undesirable.
Art methods 2:
Used cation-exchange chromatography post is cleaned into 30min with water for injection first, is then cleaned with 0.5M NaOH
90min finally rushes pillar 90min with 0.01mol/L equilibration buffer (pH6.0 ± 0.1 NaCl containing 0.1mol/L), when cleaning
Flow rate of liquid is：120ml/min；Pillar efflux is taken to carry out endotoxin and TOC detection, it is as a result undesirable.
Test example 2
Multiple real example experiment is carried out to the method for the embodiment of the present invention 1, it was demonstrated that in the conditions of the invention, use embodiment
1 method, endotoxin content, total content of organic carbon, content of microorganisms, qualification rate can achieve 99%.It tests as follows：
■ anion exchange chromatography use, which finishes, to start to clean
Flow rate of liquid is when ■ is cleaned：120ml/min
■ cleans 3900ml with water for injection
■ cleans 3900~4200ml with 0.5M NaOH NaCl containing 1M
■ cleans 3900ml with water for injection
■ cleans 3900ml with 30% isopropanol
■ cleans 3900ml with 25% acetic acid
■ cleans 3900~4200ml with water for injection
■ cleans 3900~4200ml with 0.5M NaOH NaCl containing 1M
Then ■ uses phosphate buffer（0.01MPBS（PH6.0 NaCl containing 0.1~0.2M））Rush pillar 13000ml；
After ■ is cleaned, chromatographic column is stored in 2~12 DEG C of chromatography cabinet
Wherein, the detection method of endotoxin content is as follows：
1.1 examine preceding prepare
1.1.1 prepare reagents, positive control solution, baterial endotoxin test water, sample and experiment phase before experiment starts
Close articles（Except heat source pipette tips, pipettor, alcohol, scissors, sealed membrane etc.）；
1.1.2 dry powder-shaped reagents ampoule bottle top is touched, dry powder in bottle is made to be retained in bottom of bottle portion, avoids losing when corkage
Lose powder；
1.1.3 ampoule bottle outer wall is wiped one time with 75% alcohol, after alcohol volatilization, carefully breaks ampoule bottle into two with one's hands.
1.2 sample treatment
1.2.1 it is tested using the reagents of λ=0.25EU, it is positive that sample treatment carries out positive control, test sample simultaneously
And negative control experiment, as experiment whether true foundation.
1.2.2 positive control：Reagents 1 is taken, 0.1ml baterial endotoxin test is added thereto and is redissolved with water, then plus
Enter 0.1ml bacterial endotoxin positive control solution（0.5EU/ml）；
1.2.3 negative control：Reagents 1 is taken, 0.2ml baterial endotoxin test water is added thereto.
1.2.4 test sample is positive：Reagents 1 is taken, 0.1ml purification solution sample is added thereto and redissolves, adds
0.1ml bacterial endotoxin positive control solution（0.5EU/ml）；
1.2.5 sample treatment：Reagents 1 is taken, 0.1ml baterial endotoxin test is added thereto and is redissolved with water, then plus
Enter 0.1ml sample（Concrete operations can see the table below）；
| Positive control | Negative control | Test sample is positive | Sample | |
| Baterial endotoxin test water | 0.1ml | 0.2ml | 0.1ml | |
| Positive control solution | 0.1ml | 0.1ml | ||
| Sample solution | 0.1ml | 0.1ml |
1.2.6 sample is kept the temperature：After reagents ampoule bottle after sample-adding is mixed gently, is sealed with sealed membrane, be placed on test tube
On frame；This rack for test tube is put into 37 DEG C ± 1 DEG C of constant temperature, keeps the temperature 60 ± 2min, insulating process, which should be avoided, to be shaken.
1.2.7 result is observed
1.2.7.1 after keeping the temperature, the rack for test tube for being placed with reagents ampoule bottle is taken out from constant incubator；
1.2.7.2 reagents ampoule bottle is carefully picked up from rack for test tube, slowly reverses 180 °；
If 1.2.7.3 forming gel in pipe, and gel is indeformable, is not denoted as from tube wall slippage person for the positive（+）；
1.2.7.4 not formed gel or the gel of formation be not solid, deforms and is denoted as from tube wall slippage person for feminine gender（-）；
1.2.8 inspection result determines
1.2.8.1 establishment condition is tested：Positive control pipe and the test sample positive are（+）, negative control pipe is（-）, should
Experiment can be set up.
1.2.8.2 sample cell is（-）, then show sample detection qualification（<0.25EU/ml）
1.2.8.3 if sample is（+）, take ready sample to carry out retrial 2, two results are shown（-）, then show sample
Detection is qualified（<0.25EU/ml）, otherwise show that sample detection is unqualified（>0.25EU/ml）
Wherein
The detection method of total content of organic carbon is as follows：
Pillar cleaning solution drips phosphoric acid, and whether there is or not precipitatings to generate for observation, and such as without precipitating, it is clear to measure this with total organic carbon analyzer
Washing lotion, measurement result are lower than 500ppm, then it represents that cleaning is qualified；If there is precipitating to generate, MilliQ water must be used cleaning solution dilution
Until the cleaning solution after dilution is measured the cleaning solution with total organic carbon analyzer after occurring after phosphoric acid is added dropwise without precipitating, measure
As a result it is lower than 500ppm multiplied by the data that extension rate obtains, then it represents that cleaning is qualified.
The detection method of content of microorganisms is as follows：
1.0 the method for inspection：
1.0.1 experimental implementation carries out in biological clean bench.
1.0.2 before experiment starts, 1 block of soybean casein fine jade of each placement in left and right in the working region of biological clean bench
The prefabricated plate of rouge culture medium 90mm is used for monitoring operation environment.
1.0.3 test liquid（1:10）Preparation：
1.0.3.1 water-soluble liquid test sample：Test sample 10ml is pipetted in the sterile triangular flask of 250ml with Sterile pipette
In, pH7.0 sterile NaCl-peptone buffer agent to 100ml is added（Graduation mark）, mix, as 1:10 test liquid.
1.2.8.1 water-soluble solid or semisolid test sample：Test sample 10g is weighed in the sterile triangular flask of 250ml, is added
PH7.0 sterile NaCl-peptone buffer agent to 100ml（Graduation mark）, light shaking makes to be completely dissolved, mixed with suitable method
It is even, as 1:10 test liquid.
1.0.3.2 water-insoluble test sample and the test sample of test liquid need to be prepared with specific process：It is advised according to pharmacopeia correlation
Surely it is prepared.
1.0.4 the filtering of test liquid and pad pasting：
1.0.4.1 filter bowl and filter membrane are filled：
◆ peristaltic pump：The packaging with membrane filtration cup is opened, is buckled on the buckle of peristaltic pump, cup lid is opened.
◆ microorganism detection filters system：
● alcolhol burner is lighted, cotton ball soaked in alcohol is clamped with haemostatic clamp and lights, three stainless steel stent filtering heads alcohol swab fire
Flame over-fires disinfection one by one, and flame is allowed to come into full contact with inside filtering head and edge.Sanitized puts into the cotton ball soaked in alcohol of burning
In the beaker for collecting waste.
● the sterilization packaging for opening filter core takes haemostatic clamp calcination 1min or so on alcolhol burner flame to sterilize, from packaging bag
In press from both sides out sterilizing filter element, be placed in filtering head one by one.
● it takes anodontia tweezers calcination 1min or so on alcolhol burner flame to sterilize, opens sterilised membrane filter packaging（It is careful not to
Filter membrane is encountered with hand）, clamp filter membrane and be placed on the filtering mouth of suction filtration system.
● one bag of sterile filter bowl is opened from the bottom of packaging, holds the filter mouth for being carefully buckled in stainless steel stent in the middle part of filter bowl
On, and confirm filter bowl clamping.
1.0.4.2 filter membrane is soaked
◆ filter membrane should be first soaked before water-soluble test liquid filtering, takes about 10~20ml flushing liquor（Whole filter membrane can be soaked
Subject to the amount on surface）It is poured into filter bowl along filter bowl mouth；Peristaltic pump or vacuum diaphragm pump are opened, will be closed after solution filter to the greatest extent.
◆ oils test sample, filter membrane and filter should be dried sufficiently before use, must not moisten film.
1.0.4.3 test liquid filters：
◆ 10ml test liquid (being equivalent to test sample of the every filter membrane containing 1g or 1ml), which is drawn, with 10ml Sterile pipette is added
Suitable diluent（About 100ml）In, it mixes；
◆ after poured into filter bowl along filter bowl mouth；Peristaltic pump or empty diaphragm pump are opened, will be closed after solution filter to the greatest extent.
1.0.4.4 filter membrane is rinsed：
◆ non-biocidal property product to be checked：Flushing liquor is poured into the 100ml graduation mark in filter bowl to filter bowl along filter bowl mouth, is opened compacted
Dynamic pump or vacuum diaphragm pump will close after solution filter to the greatest extent.It rinses 2 times altogether, each 100ml.
◆ biocidal property product to be checked：Its flushing dose should be confirmed by verifying, examining every time should be by authenticated flushing dose to filter
Film is rinsed, and each flushing dose of every filter membrane is 100ml, and total flushing dose must not exceed 1000ml, to avoid micro- on filter membrane
Biology is damaged.
◆ it should be noted that keeping test liquid and flushing liquor to cover entire filter membrane surface, to play the maximum of filter membrane in flushing process
Filter efficiency.
1.0.4.5 pad pasting：
◆ remove filter bowl：
● peristaltic pump：Flicking band membrane filtration cup cup body, makes filter bowl and UF membrane, removes filter bowl.
● microorganism detection filters system：Holding filter bowl edge or less lightly allows filter to tilt backwards, will from bracket
Filter bowl is removed.Anodontia tweezers are taken, cooling a moment after alcolhol burner flame sterilization（In order to avoid burning out filter membrane）, careful to clamp on filter
Filter membrane is affixed on agar medium.
◆ when pad pasting, after filter membrane one end is contacted with media surface（Bacterium is face-up）, filter membrane slowly close to media surface,
Ensure that filter membrane inner ring does not have bubble, until whole filter membrane is affixed on culture medium, then gently drives filter membrane outer ring out of with anodontia tweezers
Bubble, make whole there is no bubble formation between filter membrane and media surface.
◆ by the standard requirements of test sample, it is affixed on the prefabricated plate of soybean casein agar medium or the training of Rose Bengal Sodium agar
It supports on the prefabricated plate of base.
◆ after pad pasting, culture ware lid is covered, sample ID, lot number is recorded in culture dish bottom and is inverted in biology after the date
On clean bench.
1.0.5 the filtering of negative control and pad pasting：Diluent 10ml is taken to add after moistening film referring to 6.4.4 lower content operations
Enter suitable diluent（About 100ml）In, filtered after mixing, then with flushing liquor rinse filter membrane after pad pasting.
1.0.6 Check-Out Time：Test liquid must not exceed 1h from preparation to inspection culture medium, time is added.
1.0.7 in experimentation, at interval of 15 minutes or so, disappeared again with sterile 75% ethanol solution sprinkling hand
Poison.
1.0.8 experiment all finishes
1.0.8.1 2 pieces of prefabricated plates of soybean casein agar medium 90mm are taken, each one piece of right-hand man, the five fingers are pressed
In
5 seconds on soybean casein agar medium face, culture ware lid is covered.
1.0.8.2 the prefabricated plate of 90mm on biological clean bench is packed up, covers culture ware lid.
1.1 culture：Unless otherwise specified, it is cultivated according to following temperature and times.
1.1.1 the prefabricated plate of soybean casein agar medium is inverted in incubator, 30~35 DEG C are cultivated 5 days, by
Day point inspection.
1.1.2 the prefabricated plate of Rose Bengal Sodium agar medium is inverted in incubator, 23~28 DEG C cultivate 5 days, day by day
Point inspection.
1.2 counting：In culture period, point examines clump count day by day
If 1.2.1 asepsis growth is calculated as 0.
If 1.2.2 having 2 or 2 or more bacterium colony overlappings on plate, when distinguishable still in terms of 2 or 2 or more bacterium colonies
Number；Bacterium colony sprawling, which grows sheet of plate, to be counted, and TNTC is filled in.If there is TNTC, that is, terminate culture.
1.2.3 soybean casein agar medium is used for count of bacteria, and Rose Bengal Sodium agar medium is used for mould and ferment
Female bacterium counts；If on soybean casein agar medium on yeast and mold, Rose Bengal Sodium agar medium with thin
Bacterium, should put respectively meter yeast and mold, bacterium clump count, then by soybean casein agar medium mould and ferment
Bacterial population on female bacterium or Rose Bengal Sodium agar medium, with yeast and mold number on Rose Bengal Sodium agar medium or big
Bacterial population on beans casein agar culture medium is compared, and the clump count in the culture medium high using bacterium number is as count results.
1.3 result judgement：
1.3.1 experiment effectiveness：
1.3.1.1 negative control plate is after cultivation（Incubation time is consistent with sample）, clump count should be 0（That is < 1）.
1.3.1.2 the clump count on every filter membrane answers≤100CFU.
1.3.2 result calculates：
1.3.2.1 result report value：Point inspection most all day, point meter plate as a result, compare soybean casein agar culture medium and
Bacterium, saccharomycete clump count and the mold colony number of meter are put on Rose Bengal Sodium agar medium.Wherein the larger value conduct is taken respectively
Final bacterium colony numerical value.
1.3.2.2 calculating every 1g（ml）The micro organism quantity of test sample, formula are as follows：
Inspection product result（CFU/g（ml））The result report value of each bacterium of=test sample（CFU）÷ test sample filtration yield g（ml）×
Test sample report amount g（ml）.
Anion exchange chromatography is cleaned by above-mentioned cleaning method, so that anion exchange chromatography cleaning effect
Fruit is ensured, the endotoxin in anion exchange chromatography is eliminated, and microorganism and last time use rear remaining egg
It is white.
Specific embodiment
The present invention is further illustrated by the following examples.
Embodiment 1
Used cation-exchange chromatography post is cleaned into 30min with water for injection first, is then cleaned with 1M NaOH
90min finally rushes pillar 90min with 0.01mol/L equilibration buffer (pH6.0 ± 0.1 NaCl containing 0.1mol/L), when cleaning
Flow rate of liquid is：120ml/min；Take cleaning solution 10ml in a test tube, 2~3 drop phosphoric acid of drop, whether there is or not precipitatings to generate for observation, if
There is precipitating to generate, after occurring after must being diluted with MilliQ water until phosphoric acid is added dropwise without precipitating, total organic carbon can be transferred to
Special-purpose bottle, then three times with total organic carbon analyzer measurement, measurement result is below 500ppm three times, then it represents that cleaning is qualified；
After cleaning is qualified, anion exchange chromatography is stored in 2~12 DEG C of chromatography cabinet.
Claims (9)
1. a kind of cleaning method of cation-exchange chromatography post, which is characterized in that include the following steps：
Step 1,
By unclean cation-exchange chromatography post filler, wash with water, then cleaned with sodium hydroxide solution；
Step 2,
It is cleaned with the equilibration buffer of pH5-8, the equilibration buffer is phosphate buffer and contains NaCl；
Step 3,
The cleaning solution that step 2 obtains is taken, phosphoric acid is dripped, whether there is or not precipitatings to generate for observation, such as without precipitating, is measured with total organic carbon analyzer
The cleaning solution, measurement result are lower than 500ppm, then it represents that and cleaning is qualified,
Step 4,
If there is precipitating to generate, after occurring after step 2 must being obtained cleaning solution dilution until phosphoric acid is added dropwise with water without precipitating, will dilute
Cleaning solution afterwards measures the cleaning solution with total organic carbon analyzer, and measurement result is lower than multiplied by the data that extension rate obtains
500ppm, then it represents that cleaning is qualified；Sodium hydroxide solution described in step 1 is the sodium hydrate aqueous solution of 1-1.5M, cleaning
Time 90min or more.
2. cleaning method according to claim 1, which is characterized in that
The cation-exchange chromatography post filler is SP Sepharose Fast Flow.
3. cleaning method according to claim 1, which is characterized in that
The model of the cation-exchange chromatography post is selected from：BPG 100, BPG 500, one in XK2, XK20, XK50, XK 20
Kind.
4. cleaning method according to claim 1, which is characterized in that
The high ratio of cation-exchange chromatography post pillar diameter：(0.5-1)：1.
5. cleaning method according to claim 4, which is characterized in that
The high ratio of pillar diameter：(0.5-0.7)：1.
6. cleaning method according to claim 1, which is characterized in that
It is to clean filler in chromatographic column, cleaning solution flow velocity is 120-150ml/min.
7. cleaning method according to claim 1, which is characterized in that
Water described in step 1 is water for injection, scavenging period 30-45min.
8. cleaning method according to claim 1, which is characterized in that the sodium hydrate aqueous solution is 1.0M sodium hydroxide water
Solution, scavenging period 90-100min.
9. cleaning method according to claim 1, which is characterized in that
Equilibration buffer is 0.01mol/L phosphate buffer, pH6.0, NaCl containing 0.1-0.2M.
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4385113A (en) * | 1978-03-20 | 1983-05-24 | Nasa | Rapid, quantitative determination of bacteria in water |
| CN101224436A (en) * | 2007-09-25 | 2008-07-23 | 张剑秋 | Ion exchange resin regeneration method |
| CN101450331A (en) * | 2008-12-17 | 2009-06-10 | 牛继星 | Ion exchange resin regeneration technique capable of saving acid and alkali |
-
2014
- 2014-03-20 CN CN201410105805.2A patent/CN104922936B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4385113A (en) * | 1978-03-20 | 1983-05-24 | Nasa | Rapid, quantitative determination of bacteria in water |
| CN101224436A (en) * | 2007-09-25 | 2008-07-23 | 张剑秋 | Ion exchange resin regeneration method |
| CN101450331A (en) * | 2008-12-17 | 2009-06-10 | 牛继星 | Ion exchange resin regeneration technique capable of saving acid and alkali |
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| CN104922936A (en) | 2015-09-23 |
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Address after: 201203 No. 1 and No. 2 Juli Road, China (Shanghai) Free Trade Pilot Area, Pudong District, Shanghai Patentee after: Tian Shi Li biological medicine Limited by Share Ltd Address before: 201203 No. 280 Curie Road, Zhangjiang High-tech Park, Pudong New Area, Shanghai Patentee before: Shanghai Tasly Pharmaceutical Co., Ltd. |