CN101498832A - Cultivation viewing pool for multifunctional laser scanning co-focusing microscope - Google Patents
Cultivation viewing pool for multifunctional laser scanning co-focusing microscope Download PDFInfo
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- CN101498832A CN101498832A CNA2009100715287A CN200910071528A CN101498832A CN 101498832 A CN101498832 A CN 101498832A CN A2009100715287 A CNA2009100715287 A CN A2009100715287A CN 200910071528 A CN200910071528 A CN 200910071528A CN 101498832 A CN101498832 A CN 101498832A
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Abstract
A cultivation observation pond used on a multifunctional laser scanning confocal microscopy relates to a cultivation observation pond. The invention aims to solve the problem in the present cultivation observation pond used on a multifunctional laser scanning confocal microscopy which has the disadvantages of large difficulty, low yield, low testing precision and easy to produce syphonage. In the invention, a pond wall plate and a pond bottom plate are made into an entity; an upper terminal surface of the pond bottom plate is provided with a cultivation observation pond and two perfusion ponds; the two perfusion ponds are symmetrically arranged along an axis of the cultivation observation pond and are all communicated with the cultivation observation pond; the depths of the two perfusion ponds indicated with h are all less than the depth of the cultivation observation pond indicated with H; the distance between the bottom of the cultivation observation pond and the lower terminal surface of the pond bottom plate is between 0.1mm and 1mm. The invention is easy to process and the yield rate reaches to over 99.8 percent; the invention avoids cell culture fluid from flowing away, thus ensuring the drug concentration invariableness calculated in advance and raising the test precision; besides, the invention avoids syphonage from happening effectively.
Description
Technical field
The present invention relates to a kind of cultivation claire.
Background technology
At present, the special-purpose double dish of laser scanning co-focusing microscope that medical circle generally adopts is to be circular hole 1-1 about 10mm opening a diameter on the base plate 1, stick with glue a cover glass 2 that 0.13mm～0.17mm is thick again, form cylindrical empty pond between the wall of circular hole 1-1 and the cover glass 2 and be used for cultured cell, the fluorescence probe mark also carries out (as shown in Figure 1) in cylindrical empty pond.There is following shortcoming in above-mentioned double dish: one, because cover glass 2 is very thin and frangible, therefore bring difficulty to processing, yield rate is low; Two, the degree of depth of double dish is more shallow, and quantitatively the nutrient solution that adds very easily flows out, and then adds medicine in nutrient solution, because of nutrient solution flows out the drug concentration that calculating is good is in advance changed, thereby has reduced experimental precision; Three, the degree of depth of double dish is more shallow also can produce siphonage when perfusion.
Summary of the invention
The objective of the invention is in order to solve that the difficulty of processing that the special-purpose double dish of existing laser scanning co-focusing microscope exists is big, yield rate is low, experimental precision is low and easily to produce the problem of siphonage, and then a kind of cultivation viewing pool for multifunctional laser scanning co-focusing microscope is provided.
Technical scheme of the present invention is: cultivation viewing pool for multifunctional laser scanning co-focusing microscope comprises the pond body, described pond body is made up of pool wall plate and pond base plate, described pool wall plate and pond base plate are made one from top to bottom, pond, upper edge, the upper surface body of described pond base plate axially have a cultivation claire, pond, upper edge, the upper surface body of described pond base plate axially also have two perfusion ponds, described two perfusion ponds are symmetrical arranged along the axis of cultivating claire, described two perfusion ponds all are communicated with the cultivation claire, and the degree of depth h in two perfusion ponds is all less than the depth H of cultivating claire, and the bottom surface of described cultivation claire is 0.1mm～1mm to the lower surface of pond base plate apart from δ.
The present invention compared with prior art has following beneficial effect: the present invention is easy to processing by the die casting one-shot forming, and yield rate reaches more than 99.8%; Cultivation claire of the present invention is communicated with two perfusion ponds, can regularly or at any time carry out perfusion replacing nutrient solution to cultivating claire, liquid measure is cultivated in control, avoided the loss of nutrient solution, growth provides a good environment to cell, thereby it is constant to have guaranteed to calculate good drug concentration in advance, has improved experimental precision; Cultivation claire of the present invention in addition is communicated with the generation of also effectively having avoided siphonage with two perfusion ponds.
Description of drawings
Fig. 1 is the structural representation of the special-purpose double dish of existing laser scanning co-focusing microscope, Fig. 2 is that master of the present invention looks cut-open view, Fig. 3 is the vertical view of Fig. 2, Fig. 4 is the observation figure of acute promyelocytic leukemia NB4 cell under laser scanning co-focusing microscope that the special-purpose double dish of existing laser scanning co-focusing microscope is cultivated, Fig. 5 is the observation figure of being separated by 2 minutes with Fig. 4, Fig. 6 is the observation figure of acute promyelocytic leukemia NB4 cell under laser scanning co-focusing microscope that the present invention cultivates, and Fig. 7 is the observation figure of being separated by 2 minutes with Fig. 6.
Embodiment
Embodiment one: the cultivation viewing pool for multifunctional laser scanning co-focusing microscope of (referring to Fig. 2 and Fig. 3) present embodiment comprises pond body 3, described pond body 3 is made up of pool wall plate 4 and pond base plate 5, described pool wall plate 4 and pond base plate 5 are made one from top to bottom, claire 5-1 is cultivated in axially having of pond, upper edge, the upper surface body 3 of described pond base plate 5, pond, upper edge, the upper surface body 3 of described pond base plate 5 axially also have two perfusion pond 5-2, described two perfusion pond 5-2 are symmetrical arranged along the axis of cultivating claire 5-1, described two perfusion pond 5-2 all are communicated with cultivation claire 5-1, and the degree of depth h of two perfusion pond 5-2 is all less than the depth H of cultivating claire 5-1, and the bottom surface of described cultivation claire 5-1 is 0.1mm～1mm to the lower surface of pond base plate 5 apart from δ.
Embodiment two: the bottom surface of the cultivation claire 5-1 of (referring to Fig. 2) present embodiment is 0.13mm to the lower surface of pond base plate 5 apart from δ.So be provided with, experimental precision is higher, and the observation figure under the laser scanning co-focusing microscope is clear.Other composition is identical with embodiment one with annexation.
Embodiment three: the bottom surface of the cultivation claire 5-1 of (referring to Fig. 2) present embodiment is 0.17mm to the lower surface of pond base plate 5 apart from δ.So be provided with, experimental precision is higher, and the observation figure under the laser scanning co-focusing microscope is clear.Other composition is identical with embodiment one with annexation.
Embodiment four: scribble cell growth substrate coating 7 on the bottom surface of the cultivation claire 5-1 of (referring to Fig. 2) present embodiment.So be provided with, shortened the required time of cell attachment, impel blood primary cell and suspension cell adherent fully.Other composition is identical with embodiment one, two or three with annexation.
Embodiment five: the cell growth substrate coating 7 on the bottom surface of the cultivation claire 5-1 of (referring to Fig. 2) present embodiment is the poly-D-lysine coating.Molecular weight is that 7～30ku poly-D-lysine (poly-lysine) can make the cell attachment time foreshorten to 1 hour, and the acute isolation cell attachment time foreshortens to 30 minutes; And impel blood primary cell, suspension cell adherent fully, the adherent time of blood primary cell and suspension cell is in 10 hours.Other composition is identical with embodiment four with annexation.
Embodiment six: the cell growth substrate coating 7 on the bottom surface of the cultivation claire 5-1 of (referring to Fig. 2) present embodiment is the fibronectin coating.Fibronectin (fibronectin) can make the adherent time of attached cell foreshorten to 4 hours, and the acute isolation cell attachment time foreshortens to 30 minutes; And impel blood primary cell, suspension cell adherent fully, the adherent time of suspension cell, the adherent time of blood primary cell was in 11 hours in 12 hours.Other composition is identical with embodiment four with annexation.
Embodiment seven: the cell growth substrate coating 7 on the bottom surface of the cultivation claire 5-1 of (referring to Fig. 2) present embodiment is the glutinous protein coating of layer.The glutinous albumen of layer (laminin) can make the adherent time of attached cell foreshorten to 3.5 hours, and the acute isolation cell attachment time foreshortens to 1 hour; And impel blood primary cell, suspension cell adherent fully, the adherent time of suspension cell, the adherent time of blood primary cell was in 8 hours in 7 hours.Other composition is identical with embodiment four with annexation.
Embodiment eight: the cell growth substrate coating 7 on the bottom surface of the cultivation claire 5-1 of (referring to Fig. 2) present embodiment is a collagenic coating.Collagen can make the adherent time of attached cell foreshorten to 3.5 hours, and the acute isolation cell attachment time foreshortens to 25 minutes; And impel blood primary cell, suspension cell adherent fully, the adherent time of suspension cell, the adherent time of blood primary cell was in 9 hours in 8 hours.Other composition is identical with embodiment four with annexation.
Embodiment nine: the cell growth substrate coating 7 on the bottom surface of the cultivation claire 5-1 of (referring to Fig. 2) present embodiment is tough glutinous element coating.Tough glutinous element (tenascins) can make the adherent time of attached cell foreshorten to 5 hours, and the acute isolation cell attachment time foreshortens to 30 minutes; And impel blood primary cell, suspension cell adherent fully, the adherent time of suspension cell, the adherent time of blood primary cell was in 12 hours in 9 hours.Other composition is identical with embodiment four with annexation.
Embodiment ten: the cell growth substrate coating 7 on the bottom surface of the cultivation claire 5-1 of (referring to Fig. 2) present embodiment is the Laminin ELISA coating.Laminin ELISA (vitronectin) can make the adherent time of attached cell foreshorten to 4.5 hours, and the acute isolation cell attachment time foreshortens to 30 minutes; And impel blood primary cell, suspension cell adherent fully, the adherent time of suspension cell, the adherent time of blood primary cell was in 9 hours in 8 hours.Other composition is identical with embodiment four with annexation.
Embodiment 11: the cultivation viewing pool for multifunctional laser scanning co-focusing microscope of (referring to Fig. 2) present embodiment also increases pond lid 6, and described pond lid 6 lids are contained on the upper surface of pond body 3.So be provided with, effectively prevented the volatilization of medicine, avoided calculating good drug concentration in advance and changed because of volatilization, make experimental precision more accurate because of medicine.Other composition is identical with embodiment four with annexation.
Adopt existing double dish and the present invention under identical condition to acute promyelocytic leukemia NB4 cell, use the tyrode's solution washed twice, Fluo-3/AM with 5 μ mol/L dyes then, add 0.03% Pluronic F-127 again and in 37 ℃ of thermostatted waters are cultivated, hatch 45min, then with tyrode's solution again washed twice remove the residual Fluo-3/AM in extracellular, observe under laser scanning co-focusing microscope at last, it observes figure as shown in Figure 4 and Figure 5.Described laser scanning co-focusing microscope is produced by Japanese Olympus strain commercial firm (Olympus), and its model is Fluoview-FV300.By Fig. 4 and Fig. 5 as can be known, adopt image blurring that existing double dish obtains, and cell wherein moves obviously; By Fig. 6 and Fig. 7 as can be known, the clear picture that adopts the present invention to obtain, cell moves not obvious, and visible cell has been attached to fully to cultivate and has observed on the pool wall.
Claims (6)
1, a kind of cultivation viewing pool for multifunctional laser scanning co-focusing microscope, it comprises pond body (3), it is characterized in that: described pond body (3) is made up of pool wall plate (4) and pond base plate (5), described pool wall plate (4) and pond base plate (5) are made one from top to bottom, claire (5-1) is cultivated in axially having of pond, upper edge, the upper surface body (3) of described pond base plate (5), pond, upper edge, the upper surface body (3) of described pond base plate (5) axially also have two perfusion ponds (5-2), described two perfusion ponds (5-2) are symmetrical arranged along the axis of cultivating claire (5-1), described two perfusion ponds (5-2) all are communicated with cultivation claire (5-1), and the degree of depth h in two perfusion ponds (5-2) is all less than the depth H of cultivating claire (5-1), and the bottom surface of described cultivation claire (5-1) is 0.1mm～1mm to the lower surface of pond base plate (5) apart from δ.
2, according to the described cultivation viewing pool for multifunctional laser scanning co-focusing microscope of claim 1, it is characterized in that: the bottom surface of described cultivation claire (5-1) is 0.13mm to the lower surface of pond base plate (5) apart from δ.
3, according to the described cultivation viewing pool for multifunctional laser scanning co-focusing microscope of claim 1, it is characterized in that: the bottom surface of described cultivation claire (5-1) is 0.17mm to the lower surface of pond base plate (5) apart from δ.
4, according to claim 1,2 or 3 described cultivation viewing pool for multifunctional laser scanning co-focusing microscope, it is characterized in that: scribble cell growth substrate coating (7) on the bottom surface of described cultivation claire (5-1).
5, according to the described cultivation viewing pool for multifunctional laser scanning co-focusing microscope of claim 4, it is characterized in that: the cell growth substrate coating (7) on the bottom surface of described cultivation claire (5-1) is poly-D-lysine coating, fibronectin coating, the glutinous protein coating of layer, collagenic coating, tough glutinous element coating or Laminin ELISA coating.
6, according to the described cultivation viewing pool for multifunctional laser scanning co-focusing microscope of claim 4, it is characterized in that: described culture pond also comprises Chi Gai (6), and described Chi Gai (6) lid is contained on the upper surface of pond body (3).
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| Application Number | Priority Date | Filing Date | Title |
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| CNA2009100715287A CN101498832A (en) | 2009-03-12 | 2009-03-12 | Cultivation viewing pool for multifunctional laser scanning co-focusing microscope |
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| CNA2009100715287A CN101498832A (en) | 2009-03-12 | 2009-03-12 | Cultivation viewing pool for multifunctional laser scanning co-focusing microscope |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103823025A (en) * | 2014-03-04 | 2014-05-28 | 浙江大学宁波理工学院 | Device for screening adding or taking-out programs of antifreeze agent solution for cells |
| CN106940304A (en) * | 2016-01-04 | 2017-07-11 | 天津科技大学 | A kind of method that utilization CLSM evaluates bonded area between high yield pulp fiber |
-
2009
- 2009-03-12 CN CNA2009100715287A patent/CN101498832A/en active Pending
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103823025A (en) * | 2014-03-04 | 2014-05-28 | 浙江大学宁波理工学院 | Device for screening adding or taking-out programs of antifreeze agent solution for cells |
| CN103823025B (en) * | 2014-03-04 | 2016-02-10 | 浙江大学宁波理工学院 | Antifreeze solution for cell adds or takes out program screening plant |
| CN106940304A (en) * | 2016-01-04 | 2017-07-11 | 天津科技大学 | A kind of method that utilization CLSM evaluates bonded area between high yield pulp fiber |
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Open date: 20090805 |