CN207079243U - Disturbing flow device - Google Patents
Disturbing flow device Download PDFInfo
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- CN207079243U CN207079243U CN201721025853.6U CN201721025853U CN207079243U CN 207079243 U CN207079243 U CN 207079243U CN 201721025853 U CN201721025853 U CN 201721025853U CN 207079243 U CN207079243 U CN 207079243U
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Abstract
The utility model discloses a kind of disturbing flow device being applied in container, it includes：A plurality of baffle mechanisms, the baffle mechanism is fixedly connected by connecting rod with lower retainer ring, and a plurality of baffle mechanisms are intervally installed, the lower retainer ring is connected by rib with upper retainer ring, the side wall with the container, bottom wall swelling are adapted to respectively for fixed ring, lower retainer ring, wherein, the baffle mechanism is gradually reduced from the center line of the container to the direction of the container side wall, and the baffle mechanism forms 5~45 ° of inclination angle when the container is stood on horizontal plane with the horizontal plane.Disturbing flow device provided by the utility model can be built in the container (such as Tissue Culture Flask) of laboratory research and industrialized production, can improve the oxygen transfer efficiency in container, there is provided good ventilation and mixed effect.
Description
Technical field
A kind of disturbing flow device is the utility model is related to, can apply in the container of laboratory research and industrialized production,
To play a part of flow-disturbing.
Background technology
Tissue Culture Flask is widely used in laboratory research and industrialized production.Tissue Culture Flask can be in limited people
Under conditions of power, material resources, space, substantial amounts of experimental data is obtained.In addition, shake flat experiment result provides the basic growth of cell
Information and culture process parameter, Technical Reference is provided for mass cell culture.But traditional Tissue Culture Flask still suffer from
Lower problem：
(1) vital movement such as the growth of cell, development, division, breeding is both needed to consume substantial amounts of oxygen, participates in tricarboxylic acids and follows
Ring process.Traditional blake bottle bottleneck is small and bottom of bottle is deep and big, seriously blocks the circulation of gas.Blake bottle is on shaking table along training
Support bottle central axis to rotate, flow morphology is laminar state, is unfavorable for mixing and oxygen transmission.At present, researchers are normal
Oxygen mass transfer coefficients are improved using reducing dress liquid coefficient, putting forward high-revolving method.However, if dress liquid coefficient is too low, with cultivating
The progress of journey, the moisture in blake bottle largely evaporate, and cause the osmotic pressure of culture medium to raise, and suppress cell growth.Higher shakes
Bed rotating speed produces high shear force, causes cell damage or dissolving dead；In addition, the operating of long-time limit speed can be greatly shortened and shaken
The life-span of bed.
(2) traditional blake bottle inwall welding baffle or the concavo-convex formation baffle plate of bottle itself, this undoubtedly increases blake bottle
The difficulty of manufacturing process.The difficult point of traditional blake bottle processing is size, the location and shape that can not accurately design baffle plate,
And welding baffle component complex process in bottle.In addition, setting fixed dam in blake bottle, the difficulty of cleaning is undoubtedly also increased, it is special
Stickum caused by not being cell culture secretion is difficult to clean.
Based on disadvantages described above, the mixing that improves that (such as Tissue Culture Flask) can be built in container by how providing one kind is imitated
Fruit and turn into the direction that industry studies with the disturbing flow device of oxygen transmission efficiency.
Utility model content
Main purpose of the present utility model is to provide a kind of disturbing flow device, with overcome the deficiencies in the prior art.
To achieve these goals, the technical solution of the utility model is as follows：
The utility model embodiment provides a kind of disturbing flow device, and applied in container, it includes：
A plurality of baffle mechanisms, the baffle mechanism is fixedly connected by connecting rod with lower retainer ring, and a plurality of baffle plates
Mechanism is intervally installed, and the lower retainer ring is connected by rib with upper retainer ring, fixed ring, lower retainer ring difference
Side wall, bottom wall swelling with the container are adapted to, wherein, the baffle mechanism is from the center line of the container to the blake bottle
The direction of side wall gradually reduces, and the baffle mechanism forms 5 when the container is stood on horizontal plane with the horizontal plane
~45 ° of inclination angle.
As one of preferable preferred embodiment, described a plurality of baffle mechanisms on the bottom wall of the container in separating shape
Pluralize first recess and one second recess, first recess prolong radially from the container center lines to sidewall direction
Stretch, second recess is set around the container center lines, and first recess connects with second recess.
As one of preferable preferred embodiment, the baffle mechanism includes preceding guide face, trailing face, upper side and bottom
Face, the upper side connect to forming leading edge with the preceding guide face, and the upper side connects to forming hangover side with the trailing face,
The upper side and bottom faces connect to forming upper edge, and the preceding guide face and trailing face connect to forming inclined side, the leading edge
The first inclination angle and the second inclination angle in the same size and in opposite direction are formed with the horizontal plane respectively with hangover side.
Further, first inclination angle, the second inclination angle are 30~60 °.
Further, the angle formed between the upper side and horizontal plane is 5~45 °, and the inclined side and level
The angle formed between face is 10~175 °.
Further, the cross section of first recess is trapezoidal, in two baffle mechanisms adjacent with one first recess,
The hangover side of one baffle mechanism and the leading edge of another baffle mechanism respectively constitute the trapezoidal side, and described
The end points on the hangover side of one baffle mechanism is connected so as to be formed with the end points of the leading edge of another described baffle mechanism
The upper bottom and bottom for the trapezoidal cross-section stated.
Compared with prior art, the advantages of the utility model at least that:
Disturbing flow device provided by the utility model can be built in container (such as cell of laboratory research and industrialized production
Blake bottle) in, the oxygen transfer efficiency in container can be improved, there is provided good ventilation and mixed effect.
Disturbing flow device provided by the utility model, can when needing cleaning container independently of container (such as Tissue Culture Flask)
So that disturbing flow device to be taken out, thus it is easy to cleaning container.
Brief description of the drawings
In order to illustrate more clearly of the utility model architectural feature and technical essential, below in conjunction with the accompanying drawings and specific embodiment party
The utility model is described in detail formula.
Fig. 1 is the structural representation for the high efficiency cell container that the utility model embodiment provides；
Fig. 2 is the top view for the high efficiency cell container that the utility model embodiment provides；
Fig. 3 is the structural representation for the disturbing flow device that the utility model embodiment provides；
Fig. 4 is the structural representation for the valve protection cap that the utility model embodiment provides；
Fig. 5 is the sampling schematic diagram for the high efficiency cell blake bottle that the utility model embodiment provides；
Fig. 6 is the feed supplement schematic diagram for the high efficiency cell blake bottle that the utility model embodiment provides；
Fig. 7 is that the nutrient solution for the high efficiency cell blake bottle that the utility model embodiment provides shifts schematic diagram (force of gravity
Method)；
Fig. 8 is that the nutrient solution for the high efficiency cell blake bottle that the utility model embodiment provides shifts schematic diagram (peristaltic pump turn
Shifting method)；
Fig. 9 is in 250ml high efficiency cells blake bottle and 5L high efficiency cell blake bottles that the utility model embodiment provides
Cell number, cytoactive comparison schematic diagram.
Description of reference numerals：1- bottlenecks, 2- bottlenecks edge, 3- bottlenecks, 4- side walls, 5- side walls, 6- side walls, 7- flow-disturbings dress
Put, 7a- leading edges, 7b- hangovers side, 7c- upper edges, 7d- inclined sides, 8- external thread structures, at 9- side walls and bottom corners,
10- bottom of bottle, the recesses of 11- first, the recesses of 12- second, the upper retainer rings of 13-, 14- ribs, 15- connecting rods, retainer ring under 16-, 17-
Passage, 18- sample taps, 19- valve protection caps outside, 20- valve protection caps inner side, 21- material-feeding ports, 22- transfer ports, 23- syringes, 24- take
Sample mouth seals, probe tube in 25- bottles, 26- feed supplement pumps, the outer feed supplement pipe of 27-250ml feed supplement bottles bottle, in 28-250ml feed supplement bottle bottles
Feed supplement pipe, 29-250ml feed supplement bottles, 30- personal computers, feed supplement pipe in 31-5L blake bottle bottles, 32-5L high efficiency cell cultures
Bottle, the outer transfer pipe of 33-250ml blake bottles bottle, 34-250ml high efficiency cell blake bottles, transfer pipe in 35-250ml blake bottle bottles,
36- nutrient solution shifting pumps, 37-5L high efficiency cell blake bottles.
Embodiment
Below with reference to the accompanying drawing in the present embodiment, the technical scheme in embodiment is carried out specifically, clearly and completely to retouch
State.
As shown in figures 1-4, the disturbing flow device that the present embodiment provides is applied to high efficiency cell blake bottle, but and is limited to efficiently
Tissue Culture Flask, it can also be the container of other laboratory researches and industrialized production.Wherein, described high efficiency cell blake bottle
There is internal whorl line container and bottle cap including sealed connection, valve protection cap inner side 20, matched somebody with somebody with the external thread structure 8 of blake bottle bottleneck
Close, 19 be plane annular on the outside of valve protection cap, and there is a passage 17, one or more ports (sample tap 18, to mend at the top of valve protection cap
It is arbitrary one or more of in material mouth 21, transfer port 22).
20 on the inside of valve protection cap, an O-ring seal is provided with along at annular valve protection cap outside 19.Preferably, O-ring seal
19 inside, the internal edge adjoining annular flange (under bottle cap) of O-ring seal, loop method on the outside of external margin adjoining valve protection cap
19 are connected with O-ring seal formation on the outside of blue and valve protection cap.Preferably, O-ring seal is made up of elaxtic seal, such as
Natural rubber, buna (artificial synthetic rubber).When bottle cap and bottleneck clamping, sealing ring is located at bottleneck edge 2 and bottle
Between lid, hydraulic seal is formed.Preferably, to avoid the threaded connection between bottle cap and bottleneck excessively tight or skidding, outside valve protection cap
Long enough is answered in side 19, when bottleneck edge 2 abuts O-ring seal, the terminal abutment of bottleneck main body 3 cut-off flange.
Preferably, sealing ring includes O-ring seal or D type sealing rings.Alternative to be, sealing passes through bonding or groove
Mode connect, the size of sealing ring and the size of groove are coincide.Sealing preferably can be that individual layer sealing or bilayer are close
Envelope is multi-layer sealed, is preferably spaced at a certain distance between each layer sealing.
21 mouthfuls of described feed supplement end or transfer port 22 are connected with magnetic valve or peristaltic pump 26, peristaltic pump 36, computer control
Magnetic valve or peristaltic pump 26 processed, the switch of peristaltic pump 36 or operating, enter blake bottle to block or close feed liquid.The time of feed supplement
Can manually it be set by computer 30 with feed supplement amount.Preferably, passage 17 should (passage be most suitable straight using appropriate size
Footpath is 1.6mm), i.e., to meet that liquid can flow continually out from nutrient solution transfer port 22 and sample tap 18, avoid gas from blocking,
Liquid in bottle is avoided to spill and reduce microbiological contamination risk from passage.
In actual use, growth medium is injected into blake bottle, big unlimited bottleneck makes by the open ports of bottleneck
It must operate more simply, passage allows gas to be freely accessible to blake bottle, it is ensured that liquid continuously flows.
Alternatively, the indication wire on lid can prompt how user outwards winding bottle cap.When in blake bottle
When liquid is less than the adjoiner of bottle cap 30 and blake bottle, it is ensured that liquid stays open less than graduation mark and passage 17.
Described container further includes bottom of bottle 10, bottle main body (bottle main body includes side wall 4,5,6), bottleneck 2, bottleneck 1
With built-in disturbing flow device component 7.2.5L blake bottles base diameter is 50mm, a height of 28cm, bottleneck long 75mm, a diameter of
75mm.Preferably, blake bottle is made up of glass, polymeric material, such as polypropylene.Preferably, container can select injection
Shaping or the mode of blow molding, and can abandon, reduce the pollution for recycling and bringing.
Described container can also be in conical, cylindrical, spherical, the square bodily form, cuboid, but not limited to this
Other shapes.Preferably, container is in cone, by smooth between the bottom wall 10 and side wall 6 of container and between side wall 4,5,6
The circle turning of transition or cambered surface connection, bottle bottom wall 10 is slightly raised, when blake bottle liquid amount than it is relatively low when, the fluid in blake bottle
It can be flowed along the corner of bottle bottom wall 10.The side wall 6 of bottle and blake bottle centerline axis parallel；Side wall 5 favours center
Axis, angle of inclination are 1~80 °, it is preferred that angle of inclination is 5~45 °；Side wall 4 also favours central axis, angle of inclination
For 1~80 °, it is preferred that angle of inclination is 5~45 °.The side-walls of container may also set up liquid level line, and indicate corresponding culture
Bottle volume.
Described bottleneck 2 is located at the end of bottle side wall 4, in cylindrical, square, but is also not necessarily limited to other shapes, preferably
Be cylinder.The edge of bottleneck 3 is provided with external thread structure 8.External thread structure 8 can be single line, a plurality of line, can also
For continuous lines or interrupt line, preferably a plurality of interrupt line.The form of screw thread is not limited to latch, flange connection and other connections
Mode.Flange is located at the position below screw thread, and extends to outside bottleneck, and bottleneck 2 is small enough in one-handed performance.Bottleneck 2
Diameter and its length can change, and mark can print to turning 9 or blake bottle side wall 4,5,6.
Such as Fig. 1, in preferred example, blake bottle includes bottom of bottle 10, several disturbing flow devices 7 built in blake bottle.It is preferred that
, the bottom wall 10 of blake bottle is connected with blake bottle side wall 4,5,6 by way of circle turning；But alternative, blake bottle side
Wall 4,5,6 is tapered, with one or more gradients close to central axis.Blake bottle side wall 4,5,6 connects bottleneck 2.The side of bottleneck 2
There is one to open wide bottleneck 1 at edge.Side wall 6 and centerline axis parallel, side wall 5 slowly favour central axis, and side wall 5 rapidly favours
Central axis.With the change of the structure snd size of disturbing flow device 7, the outer wall dimension for cultivating bottle container also changes therewith.
Disturbing flow device 7 can use the elastic material (such as silica gel or rubber) with auto-reset function, can also use without bullet
Property stainless steel or hard plastic material (such as makrolon, polyester, fluoropolymer, acrylic acid, PET, polyvinyl chloride,
Any one in ABS, PC, PS, GAG, acrylic and polyolefin or two or more combinations).Above-mentioned material should be resistant to not
High-temperature sterilization processing of 121 DEG C less than the second recess, γ ray sterilization, β ray sterilizings, the processing of chemical reagent sterilization.
Disturbing flow device include upper retainer ring 13, lower retainer ring 16, rib 14, baffle plate, connecting rod 15, upper and lower retainer ring 13,
16 are linked into an integrated entity by rib 14, upper retainer ring 13 and lower retainer ring 16 side wall 4,5,6 and bottom wall 10 with blake bottle respectively
Adaptation, baffle plate are connected by connecting rod 15 with lower retainer ring 16, and in the circular space of the closing of lower retainer ring 16.
Disturbing flow device 7 plays a part of ventilation mixing, but not damaged cell.The design of disturbing flow device 7 should reduce shearing
Power, strengthen mixed effect.Therefore, when using blake bottle, speed setting is particular value, built-in disturbing flow device, can be obtained preferably
Admixture, less shearing force, avoid the breakage of cell membrane.
In instantiation, the bottle bottom wall 10 of pie is divided into the first recess 11, the second recess by built-in disturbing flow device 7
12, the first recess 11, the second recess 12 is stretched out by center line.Baffle plate be shaped as rectangle, parallelogram, square,
One or more in trapezoidal, triangle, preferably rectangle.The number of baffle plate can change, preferably even number.As Fig. 2-
Blake bottle bottom is divided into six recesses by six disturbing flow devices 7 shown in 3, the angle between adjacent two the first recesses 11
For 60 °.Certainly, it is also feasible using four disturbing flow devices 7, blake bottle bottom can be divided into four by four stream devices 7
First recess 11, the angle between adjacent two the first recesses 11 is 90 °.
Blake bottle continuously rotates along a direction, and preferably clockwise, each baffle plate of disturbing flow device 7 is along same
Sample prescription is to rotation.In actual moving process, equipped with corresponding equipment, the direction rotated is controlled by operator.Each flow-disturbing dress
7 baffle plate is put to be made up of preceding guide face, trailing face, upper side and bottom surface, any two of which homogeneously connects between face, it is described on
Portion face connects to forming leading edge 7a with the preceding guide face, and the upper side connects to forming the side 7b that trails with the trailing face, leading
While 7a and one trail while 7b respectively the slanted angle between horizontal plane be 30~60 ° (bottom wall 10 of blake bottle positioned at level
On face), more excellent, slanted angle is 30~40 °.The rotary speed of blake bottle is 20~300rpm, it is preferred that rotary speed is
80~200rpm.Certainly, higher rotary speed can be also used, but angle angle need to be adjusted, to avoid shearing from causing cell to break
Damage.Therefore, the design of disturbing flow device 10 need to consider rotary speed, to maintain low-shearing force, to avoid cell damage.
Leading edge 7a and hangover side 7b extend to center line, radially distribute.Center section 7c is generally planar, but
Can be slightly convex.Preferably, the bottle bottom wall 10 of pie is divided into the first recess 11, the second recess 12 by disturbing flow device 7, but
Shape can change.When culture bottle container is rotated with certain speed, the inclination of baffle plate leading edge 7a and hangover side 7b and horizontal plane is pressed from both sides
Angle can avoid cell damage.After cell infection, cell is collected.Stirring slightly influences ventilation, because the formation of bubble influences carefully
The breakage of cell wall.If speed of agitator increases, in order to reduce shearing force, angle may reduce.
Interval between each baffle plate of disturbing flow device 7 also influences the ventilation and stirring of fluid.First recess 11 it is transversal
Face is trapezoidal, in two baffle plates adjacent with one first recess 11, the hangover side 7b of baffle plate and with before another baffle plate
Guide margin 7a respectively constitutes the side of described trapezoidal cross-section, and the end points on the hangover side of a baffle plate is leading with another baffle plate
The end points on side is connected so as to form described trapezoidal upper bottom and bottom, and nature of going to the bottom is located at the bottom wall 10 of bottle.Preferably,
Leading edge 7a and hangover side 7b are tilted with same degree, but in the opposite direction, so that fluid lightly flows through leading edge 7a, are bypassed
Upper edge 7c, through side 7b outflows of trailing.Fluid bypasses the first recess 11, reaches next leading edge 7a, flows through next top
Side 7c, next first recess 11 is flowed out to through next hangover side 7b, is circulated with this, the lumpy fluid caused by
Flowing, beneficial to inflation, and will not damaged cell membrane.
Blake bottle rotates around center line, and the fluid velocity of center line is minimum.With from increase, fluid with a distance from central axis
Speed linearity increase.Frictional force between fluid and culture bottle container can avoid fluid from being rotated with a certain speed.(the radiation of turning 9
Shape farthest) place fluid-flow rate be more than centerline fluid-flow rate.
At neighbouring turning 9, the flowing velocity at disturbing flow device 7 is minimum；Flowing at centerline, disturbing flow device 7
Speed highest.Preferably, the disturbing flow device at neighbouring turning 9 successfully incorporates bottom wall 10 and turning 9.Preferably, flow-disturbing fills
It is center line to the 1/3 of turning outer peripheral lines distance to put the distance between 7 and turning 9 D.In instantiation, disturbing flow device is with turning
The distance between angle D about 25mm.
When blake bottle is rotated with 80~180rpm rotary speeies, the upper side of disturbing flow device favours 5~12 ° of horizontal plane,
In instantiation, preferable angle of inclination is 9 °, and preferable rotary speed is 80~100rpm.
Neighbouring centerline, the flowing velocity highest of disturbing flow device 7.The distance between disturbing flow device 7 and central axis are
The 1/8 of container diameter.Disturbing flow device 7 is tilted to the direction away from center line, and about 20~50 ° of folder is formed with horizontal plane
Angle, it is preferred that angle is 38 °.Preferably, because right angle mode connection shearing force it is big, thus avoid inclined side 7d, upper edge 7c,
Connected between leading edge 7a, hangover side 7b with right angle mode and then to avoid cellular damage, in design typically to justify turning
Mode connects.
In an instantiation, disturbing flow device 7 includes six baffle plates, spacing about 5~8mm between each baffle plate.Training
Bottle radius 2/3, neighbouring corner 9 are supported, upper edge 7c merges with bottle bottom wall 10.Six cheese disturbing flow devices 7 are divided into six
The first recess of rectangle 11, and form second recess of recess 12 in centerline.
When in use, blake bottle is equipped with the growth medium of 1/3 volume, is totally submerged disturbing flow device 7.Preferably, edge
Central line measurement, disturbing flow device 7 and the height of horizontal plane bottom are about the 1/3 of fluid level.More optimizedly, disturbing flow device
Highly it is about the 1/5~1/10 of fluid level.
Blake bottle is fixed on shaking table.Side wall 4, side wall 45 are consistent with the central shaft of fixture, are revolved with 80~180rpm speed
Turn.Fluid wave flows through disturbing flow device 7 and plays a part of stirring, mixing, inflation, avoids cell membrane damaged.
As shown in figure 5, the sampling method of the high efficiency cell blake bottle 34,32 of built-in disturbing flow device 7 is as follows：
1) the sample tap sealing 24 at sampling port 18 is outwarded winding；
2) sampled after connecting syringe 23 and sampling port 18, probe tube 25 connects with sampling port 18 wherein in bottle；
3) the sample tap sealing 24 at sampling port 18 is screwed.
As shown in fig. 6, following (the high efficiency cell culture of the feed process of the high efficiency cell blake bottle 32 of built-in disturbing flow device 7
Bottle 34 is identical)：
1) by feed supplement pipe 31 in the 5L blake bottle bottles in high efficiency cell blake bottle 32 and the feed supplement silica gel of 250ml feed supplements bottle 29
27 sterile welding of pipe；
2) feed supplement silicone tube 27 is arranged in the pump head of feed supplement pump 26, and feed supplement silicone tube 27 and 250ml feed supplement bottle bottles
Interior feed supplement pipe 28 connects；
3) rotating speed and the opening/closing time of time or magnetic valve of feed supplement pump 26 are adjusted, and opens feed supplement valve, according to cell
Upgrowth situation starts feed supplement.
Nutrient solution in the high efficiency cell blake bottle 34 of built-in disturbing flow device 7 is transferred to another high efficiency cell blake bottle 37
Method include following methods：
A) force of gravity method (as shown in Figure 7)
A1) transfer pipe in the 250ml blake bottle bottles in Tissue Culture Flask 34 is connected with transfer silicone tube 33, and to turn
Move silicone tube 33 and the 37 sterile welding of high efficiency cell blake bottle；
A2) Tissue Culture Flask 34 is inverted or adjusted to suitable position；
A3 liquid transfer valve) is opened, nutrient solution is transferred to completely to efficient Tissue Culture Flask 37.
B) peristaltic pump transfer method (as shown in Figure 8)
B1) transfer pipe in the 250ml blake bottle bottles in Tissue Culture Flask 34 is connected with transfer silicone tube 33, and to turn
Move silicone tube 33 and the 37 sterile welding of high efficiency cell blake bottle；
B2 silicone tube 33) will be shifted to be arranged in the pump head of nutrient solution shifting pump 36；
B3) rotating speed and the time of nutrient solution shifting pump 36 are set, and open liquid transfer valve, nutrient solution is transferred to completely
To efficient Tissue Culture Flask 37.
Below in conjunction with specific embodiment and accompanying drawing, the utility model is described in more detail.
Embodiment 1
To evaluate the efficient culture performance of high efficiency cell blake bottle, dress liquid coefficient, (contamination rate, single take operability
Sample is time-consuming, single inoculation transfer is time-consuming), PK-15 cells and Chinese hamster ovary celI are cultivated, respectively 50 bottles of the culture of every kind of cell, culture
Condition is as follows：
The condition of culture of PK-15 cells：(1) high efficiency cell blake bottle：5L is seeded to from 250ml high efficiency cells blake bottle 34
High efficiency cell blake bottle 32, inoculum density are 0.6 × 106Cells/ml, 2g/L the mounting medium (Cytodex- of GE companies production
1), 37 DEG C, 5%CO2Culture.Flowed every 1d using feed supplement pump 26 plus add supplemented medium.When culture is to 4d, collect thin
Born of the same parents.(2) regular growth blake bottle：5L regular growth blake bottles are seeded to from 250ml regular growth blake bottles, inoculum density is
0.6×106Cells/ml, 2g/L mounting medium (Cytodex-1 of GE companies production), 37 DEG C, 5%CO2Culture.Every 1d profits
Manually add supplemented medium.When culture is to 4d, cell is collected.
The condition of culture of Chinese hamster ovary celI：(1) high efficiency cell blake bottle：5L height is seeded to from 250ml high efficiency cells blake bottle 34
Tissue Culture Flask 32 is imitated, inoculum density is 0.8 × 106Cells/ml, 37 DEG C, 5%CO2Culture.When glucose exhausts, feed supplement
Pump 26 adds supplemented medium and glucose.When culture is to 5d, cultivation temperature is adjusted to 32 DEG C.When culture is to 14d, collect thin
Born of the same parents.(2) regular growth blake bottle：5L regular growth blake bottles are seeded to from 250ml regular growth blake bottles, inoculum density is
0.8×106Cells/ml, 37 DEG C, 5%CO2Culture.When glucose exhausts, supplemented medium and glucose are manually added.Training
When supporting to 5d, cultivation temperature is adjusted to 32 DEG C.When culture is to 14d, cell is collected.
When sampling and inoculation transfer, calculating single sampling is time-consuming and single inoculation takes；At the end of culture, it is measured by sampling most
Whole average cell number and observation microbiological contamination situation result simultaneously calculate contamination rate.
Sample 1000rpm centrifuges 15min, discards supernatant, adds 0.1% crystal violet dye liquor, 37 DEG C of water-bath 1h, using blood
Cell number in ball count plate determination sample.And cell microbiological contamination situation is observed, the calculation formula of contamination rate is：Contamination rate (%)=dye
Bacterium bottle number/(non-microbiological contamination bottle number+microbiological contamination bottle number).
Culture performance and the operability rate of exchange of the high efficiency cell blake bottle of table 1 with regular growth blake bottle
The PK-15 cells of high-efficient culture bottle culture and the cell density ratio cellar culture of Chinese hamster ovary celI are up to 53.6% respectively
With 37.8%.High-efficient culture bottle not only can guarantee that high-density cell growth, moreover it is possible to increase the liquid amount of shaking flask, the training of 5L high efficiency cells
The dress liquid coefficient for supporting bottle may be up to 60%, and the dress liquid coefficient of cellar culture bottle is only 30%.High-efficient culture bottle and cellar culture
The mode of off normal sampling and in-situ sampling is respectively adopted in bottle, and single sampling takes 6.9~7.6min and 2.1~2.2min respectively.
High-efficient culture bottle is inoculated with by the way of the transfer of feed supplement pump, the half of the time-consuming only cellar culture bottle inoculation time of single inoculation,
And it largely avoided microbiological contamination.
Embodiment 2
Investigate amplification performance of the HEK-239 cells in high efficiency cell blake bottle.The condition of culture of cell：From 250ml height
Effect Tissue Culture Flask 34 is seeded to 5L high efficiency cells blake bottle 32, and inoculum density is 0.5 × 106Cells/ml, 37 DEG C, 5%CO2
Culture.Supplemented medium is added every 1d.When culture is to 7d, cell is collected.Cell number and cytoactive are measured by sampling daily.
The method that cytoactive detection uses propidium iodide (PI) dyeing.When frustule dyes, PI storages are added into sample
(0.2 μm of membrane filtration, is preserved standby liquid at 4 DEG C, and ultimate density is 10 μm of ol/L, and 20min is incubated under dark room temperature condition.
PI dyestuffs can not mark living cells, and dead cell intracellular PI dyestuffs are launched red fluorescence after 488nm laser excitations, be led to by FL2
Road (585nm) receives, and FL2 passages at least detect 10000 cells.The analysis flow velocity of sample is 60 μ l/min, loading volume
For 10 μ l.
As shown in Figure 9, the cell growth variation tendency of HEK-239 cells is similar in 250ml with 5L high-efficient culture bottles, just
Phase is slow-growing, subsequently into fast growing period, tends to stable when cultivating 6d, when cultivating 7d cell number reach 4.6 ×
106cells/ml.The result shows that HEK-239 cells obtain basically identical growth effect in 250ml and 5L high-efficient culture bottles
Fruit.
Above-mentioned embodiment, only illustrate technical concept and architectural feature of the present utility model, it is therefore intended that allow ripe
Knowing the stakeholder of technique can implement according to this, but above said content is not intended to limit the scope of protection of the utility model,
Every any equivalent change or modification made according to Spirit Essence of the present utility model, all should fall into protection of the present utility model
Within the scope of.
Claims (6)
- A kind of 1. disturbing flow device, applied in container, it is characterised in that including：A plurality of baffle mechanisms, the baffle mechanism is fixedly connected by connecting rod with lower retainer ring, and a plurality of baffle mechanisms Be intervally installed, the lower retainer ring is connected by rib with upper retainer ring, fixed ring, lower retainer ring respectively with institute Side wall, the bottom wall swelling adaptation of container are stated, wherein, the baffle mechanism is from the center line of the container to the container side wall Direction gradually reduces, and the baffle mechanism forms 5~45 ° when the container is stood on horizontal plane with the horizontal plane Inclination angle.
- 2. disturbing flow device according to claim 1, it is characterised in that：Described a plurality of baffle mechanisms are in the container Be separated to form a plurality of first recesses and one second recess on bottom wall, first recess radially from the container center lines to Sidewall direction extends, and second recess is set around the container center lines, and first recess connects with second recess It is logical.
- 3. disturbing flow device according to claim 2, it is characterised in that：The baffle mechanism include preceding guide face, trailing face, on Portion face and bottom faces, the upper side connect to forming leading edge with the preceding guide face, and the upper side connects with the trailing face Hangover side is formed, the upper side and bottom faces connect to forming upper edge, and the preceding guide face and trailing face connect to forming inclined side, The leading edge and hangover side form in the same size and the first inclination angle and second in opposite direction with the horizontal plane and inclined respectively Oblique angle.
- 4. disturbing flow device according to claim 3, it is characterised in that：First inclination angle, the second inclination angle be 30~ 60°。
- 5. disturbing flow device according to claim 3, it is characterised in that：The angle formed between the upper side and horizontal plane For 5~45 °, and the angle formed between the inclined side and horizontal plane is 10~175 °.
- 6. disturbing flow device according to claim 2, it is characterised in that：The cross section of first recess is trapezoidal, with one In two adjacent baffle mechanisms of first recess, the hangover side of a baffle mechanism is distinguished with the leading edge of another baffle mechanism Form the trapezoidal side, and the end points on the hangover side of described baffle mechanism and another described baffle mechanism The end points of leading edge connect so as to form the upper bottom and bottom of described trapezoidal cross-section.
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| CN201721025853.6U CN207079243U (en) | 2017-08-16 | 2017-08-16 | Disturbing flow device |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109401960A (en) * | 2017-08-16 | 2019-03-01 | 苏州米迪生物技术有限公司 | The high efficiency cell culture bottle of built-in disturbing flow device |
| US20210189311A1 (en) * | 2018-05-16 | 2021-06-24 | aquila biolabs GmbH | Device and method for culturing cells in a shaking operation |
-
2017
- 2017-08-16 CN CN201721025853.6U patent/CN207079243U/en active Active
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109401960A (en) * | 2017-08-16 | 2019-03-01 | 苏州米迪生物技术有限公司 | The high efficiency cell culture bottle of built-in disturbing flow device |
| US20210189311A1 (en) * | 2018-05-16 | 2021-06-24 | aquila biolabs GmbH | Device and method for culturing cells in a shaking operation |
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