CN2748466Y - Integral system of cell culturing and product reclaiming - Google Patents
Integral system of cell culturing and product reclaiming Download PDFInfo
- Publication number
- CN2748466Y CN2748466Y CN 200420115323 CN200420115323U CN2748466Y CN 2748466 Y CN2748466 Y CN 2748466Y CN 200420115323 CN200420115323 CN 200420115323 CN 200420115323 U CN200420115323 U CN 200420115323U CN 2748466 Y CN2748466 Y CN 2748466Y
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- cell cultures
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- liquid storage
- cell culturing
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Abstract
The utility model relates to an integral system of cell culturing and product reclaiming, which comprises a cell culturing tank connected with a liquid tube. The cell culturing tank is connected into a liquid storage tank through a discharging tube, and then ordinally connected to a chromatographic column and a waste liquid tank through the discharging tube. Peristaltic pumps are arranged between the cell culturing tank and the liquid storage tank as well as between the liquid storage tank and the chromatographic column, and the peristaltic pumps controlled by a pH automatically regulating device can feed acid and alkali. The entry end of the alkaline peristaltic pump is externally connected with alkali liquor, the entry end of the acidification peristaltic pump is externally connected with acid liquor, and the exit ends of the two peristaltic pumps are accessed into the liquid storage tank; a stirrer is arranged in the liquid storage tank; the pH electrode assembly of the pH automatically regulating device is immersed into the culturing supernatant in the liquid storage tank. The pH automatically regulating device is composed of an acidimeter and an SCM real-time controlling system for timely control. The integral system of cell culturing and product reclaiming is applicable to the reclaiming of most cell product of animal engineering. Compared with the respective system of cell culturing and product reclaiming, the utility model reduces a plurality of separation devices and manpower, and also reduces the production cost.
Description
Technical field
The utility model relates to the processing unit that is used for animal engineering cell product large-scale recovery, refers to the integral system that a kind of cell cultures and product reclaim especially.
Background technology
As some scientists prophesy, 21 century is bio-science century, life science combines with other subjects just with unprecedented speed develop rapidly.Along with development of biology, the industrialization of increasing gene recombination product has produced and will continue generation huge social benefit and considerable economic.Biotechnology downstream technology is the gordian technique of biotechnology industry, and it comprises the production preparation of recovery, purifying products, product of Bio-engineering Products and production quality control or the like.The traditional recovery of product in the past technology all is that cell culture fluid or cell homogenates are removed the concentrated back of solid formation branch upward chromatographic column or direct upper prop, and product is adsorbed onto on the chromatographic column, elutes then, carry out consummateization with the high efficiency chromatography method again.Preparation is feasible for small-scale production for this, cultivates but can not satisfy the extensive perfusion continuously of Mammals engineering cell--produce the scale operation of several hectolitres, a few kilolitre cells and supernatant every day.
In order to reduce the production stage of Mammals engineering cell product, reduce production costs, American-European minority developed country is from late 1980s just the begin one's study large scale culturing of Mammals engineering cell and the integral process that product reclaims.Britain's Birmingham university research the integrated apparatus of one cover mouse hybridoma cell cultured continuously and monoclonal antibody recovery, U.S. expert has also reported a biotechnology device, can be with a large amount of cultivations of engineering cell and disposable the finishing of purifying of recombinant protein.They are that the antigen with specific monoclonal antibody is coupled on the adsorption medium with chemical process, making the affine fluidised bed chromatography of antibody annotates, affinity column is directly connected to the exit end of the continuous perfusion culture tank of hybridoma (bio-reactor), cells and supernatant is through filter screen, the fluidized-bed affinity chromatographic column of flowing through, monoclonal antibody is adsorbed onto on the chromatographic column, waste liquid flows in the waste liquid barrel from the exit end of chromatographic column, regularly affinity chromatographic column is switched then and get off to carry out wash-out, the monoclonal antibody of collection carry out consummateization with the high efficiency chromatography method again.But, this production technique only is suitable for reclaiming products such as monoclonal antibody and specific antigen, do not reclaim other most protein or other biomacromolecule and be suitable for, because most protein, nucleic acid adopt ion-exchange chromatography to reclaim more convenient, cells and supernatant is before last chromatographic column, must adjust pH value could upper prop, otherwise product not can be incorporated on the chromatographic column, also just can't realize the integrated of cell cultures technology and product recovery technology.In addition, the antibody affinity chromatographic column is not easy to sterilization, the antibody polluted product that can come off gradually in use, to satisfy simultaneously the scale operation needs prepare a large amount of affinity antibodies again needs drop into some force-summing devices.
Summary of the invention
The purpose of this utility model provides the integral system of a kind of cell cultures and product recovery, it reclaims protein or other biomacromolecule product applicable to ion exchange chromatography, realizes the integrated and scale operation that the continuous perfusion of engineering cell is cultivated and product reclaims.
For achieving the above object, the utility model is taked following design: the integral system that a kind of cell cultures and product reclaim, comprise the cell cultures jar that connects liquid-inlet pipe, the cell cultures jar is connected in the container for storing liquid by liquid discharge pipe, be connected to chromatographic column and waste liquid tank by liquid discharge pipe successively again, at cell cultures jar and container for storing liquid, be provided with peristaltic pump between container for storing liquid and chromatographic column, if be controlled by-the pH device for automatically regulating add acid, the peristaltic pump that adds alkali, add the external alkali lye of alkali peristaltic pump inlet end, add the external acid solution of sour peristaltic pump inlet end, the exit end of two peristaltic pumps all inserts in the container for storing liquid;
The pH combined electrode of described pH device for automatically regulating immerses in the interior culture supernatant of container for storing liquid.
Described pH device for automatically regulating is connected and composed by the micro-chip real-time control system of acidometer and controlling acidity meter.
In liquid storage is irritated, an agitator is installed.
The utility model has the advantages that:
(1) this device is applicable to the recovery of most of animal engineering cell products:
(2) with cell cultures with reclaim product respectively and compare, the present invention has reduced part separating device and manpower, can reduce production costs.
Description of drawings
Fig. 1 constitutes synoptic diagram for the utility model system
Fig. 2 is the utility model pH device for automatically regulating principle of compositionality block diagram
Embodiment
As shown in Figure 1, the integral system that the utility model cell cultures and product reclaim, comprise the cell cultures jar 2 that connects liquid-inlet pipe 1, the cell cultures jar is connected in the container for storing liquid 3 by liquid discharge pipe, be connected to chromatographic column 4 and waste liquid tank 5 by liquid discharge pipe successively again, between cell cultures jar and container for storing liquid, container for storing liquid and chromatographic column, be provided with peristaltic pump 6, other establishes and is controlled by-the adding acid, add the peristaltic pump 7,8 of alkali of pH device for automatically regulating, add the external alkali lye of alkali peristaltic pump 8 inlet ends, add the external acid solution of sour peristaltic pump 7 inlet ends, the exit end of two peristaltic pumps all inserts in the container for storing liquid;
The pH combined electrode of described pH device for automatically regulating 9 immerses in the interior culture supernatant of container for storing liquid.
Described chromatographic column can adopt online wash-out (Streamline) cationic exchange expanding bed chromatographic column.
Referring to Fig. 2, described pH device for automatically regulating 9 is DpH87 acidometer (Military Medical Science Institute's product) is linked to each other with 8031 micro-chip real-time control systems and to control in good time.
Described 8031 micro-chip real-time control systems comprise central processing unit 8031CPU, a 2764EPROM (system program holder), 2817A EEPROM (electric sassafras holder, the permanent programming file data of preserving), ADC0809 (8 8 road modulus conversion chips are used for the digitizing conversion of pH glass electrode signal and temperature signal); A slice programmable keyboard display interface device-8279 (be used for button and show the man-machine dialogue operation); Also have two-way bidirectional thyristor non-contact switch circuit (adopting light isolated controlling module) in addition, be used to control two peristaltic pumps and add acid or add alkali.The pH electrode of acidometer and-temperature sensor connects the modulus conversion chip input terminus of micro-chip real-time control system respectively through amplifier; Two control output ends of micro-chip real-time control system are respectively through two peristaltic pumps that add acid, add alkali of two-way bidirectional thyristor non-contact switch circuit control.
This device is applicable to the recovery of most of animal engineering cell products, and can be used for the large-scale recovery preparation of animal engineering cell product.
Use the continuous perfusion cultivation of integral system device preparation engineering cell of the utility model cell cultures and product recovery and the removal process of recombinant human urokinase zymogen (rhpro-UK) with following specific embodiment explanation:
(1) target protein is cultivated and reclaimed to engineering cell: the recovery of CHO engineering cell is cultivated in square vase, blender jar, rolling bottle successively, treat that engineering cell density reaches 1 * 10
6Change over to behind cell/ml in 5 liters of bio-reactors of B Blang (B.BRAUN) and cultivate, treat that cell density reaches 5 * 10
6Cell/ml changes 30 liters of bioreactor culture of B Blang (B.BRAUN) again over to, will press 30 liters of bio-reactors of Fig. 1 and B Blang (B.BRAUN) through the integrated apparatus of sterilization then and be connected, and chromatographic column is got up with level pad balance and expansion in advance.When cell culture fluid flows into container for storing liquid and reaches 2/3 volume, with the pH adjusting and control instrument with add the power supply opening of soda acid peristaltic pump and the pH value is set between 5～6, open online wash-out cationic exchange (streamliane SP) expanding bed chromatographic column feed liquor peristaltic pump power switch then and flow velocity is transferred to 40～50ml/ minute (daytime upper prop, stop night), changed an online wash-out cation-exchange chromatography post every 3～4 days.
(2) wash-out of target protein: the chromatographic column that is adsorbed with target protein switch get off after, the post lower end connects a peristaltic pump, the upper end is connected (wavelength 280nm) with UV-detector, with level pad (0.01mol/L phosphate buffered saline buffer, contain 0.1mol/L NaCl, pH 5～6) residual cell culture fluid is rinsed well, then the post bed is compressed to original volume, use elution buffer (0.01mol/L phosphate buffered saline buffer again, contain 1mol/L NaCl, pH 7.0) the wash-out target protein, collect eluted protein peak effluent liquid, obtain the thick product of recombinant human urine and proenzyme (rhpro-UK).
(3) consummateization of target protein: (contain 0.2mol/L NaCl with the 0.01mol/L phosphate buffered saline buffer, pH7.0) the about column volume of balance polyphenyl alkene acid amides (Sephacryl S-200HR) gel chromatographic columns, with thick product upper prop, continue drip washing with same buffer, collect albumen main peak elution fraction, purity reaches 99.98% (HPLC analysis), the rate of recovery 72.2%.
With cell cultures with reclaim product respectively and compare, the present invention has reduced part separating device and manpower, can reduce production costs.
Claims (5)
1, the integral system of a kind of cell cultures and product recovery, comprise the cell cultures jar that connects liquid-inlet pipe, the cell cultures jar is connected in the container for storing liquid by liquid discharge pipe, be connected to chromatographic column and waste liquid tank by liquid discharge pipe successively again, between cell cultures jar and container for storing liquid, container for storing liquid and chromatographic column, be provided with peristaltic pump, it is characterized in that:
If be controlled by adding acid, adding the peristaltic pump of alkali of a pH device for automatically regulating, add the external alkali lye of alkali peristaltic pump inlet end, add the external acid solution of sour peristaltic pump inlet end, the exit end of two peristaltic pumps all inserts in the container for storing liquid;
The pH combined electrode of described pH device for automatically regulating immerses in the interior culture supernatant of container for storing liquid.
2, the integral system of cell cultures according to claim 1 and product recovery is characterized in that: described pH device for automatically regulating is connected and composed by the micro-chip real-time control system of acidometer with control in good time.
3, the integral system of cell cultures according to claim 2 and product recovery is characterized in that described micro-chip real-time control system comprises:
One central processor CPU,
One system program holder EPROM,
The one permanent electric sassafras holder EEPROM that preserves the programming file data,
One is used for the modulus conversion chip of the digitizing conversion of pH glass electrode signal and temperature signal,
A slice programmable keyboard display interface device;
The pH electrode of acidometer and a temperature sensor connect the modulus conversion chip input terminus of micro-chip real-time control system respectively through amplifier;
Two control output ends of micro-chip real-time control system are respectively through two peristaltic pumps that add acid, add alkali of two-way bidirectional thyristor non-contact switch circuit control.
4, the integral system of cell cultures according to claim 2 and product recovery, it is characterized in that: described acidometer adopts the DPH87 acidometer.
5, the integral system of cell cultures according to claim 1 and product recovery is characterized in that: an agitator is installed in described liquid storage is irritated.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 200420115323 CN2748466Y (en) | 2004-11-22 | 2004-11-22 | Integral system of cell culturing and product reclaiming |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 200420115323 CN2748466Y (en) | 2004-11-22 | 2004-11-22 | Integral system of cell culturing and product reclaiming |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN2748466Y true CN2748466Y (en) | 2005-12-28 |
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| Application Number | Title | Priority Date | Filing Date |
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| CN 200420115323 Expired - Lifetime CN2748466Y (en) | 2004-11-22 | 2004-11-22 | Integral system of cell culturing and product reclaiming |
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| CN (1) | CN2748466Y (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100370022C (en) * | 2004-11-22 | 2008-02-20 | 中国人民解放军军事医学科学院生物工程研究所 | Technology for culturing cell and recovering products, its integrating system thereof |
-
2004
- 2004-11-22 CN CN 200420115323 patent/CN2748466Y/en not_active Expired - Lifetime
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100370022C (en) * | 2004-11-22 | 2008-02-20 | 中国人民解放军军事医学科学院生物工程研究所 | Technology for culturing cell and recovering products, its integrating system thereof |
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Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| AV01 | Patent right actively abandoned |
Effective date of abandoning: 20080220 |
|
| AV01 | Patent right actively abandoned |
Effective date of abandoning: 20080220 |
|
| C25 | Abandonment of patent right or utility model to avoid double patenting |