细胞世界

因为不知道你们具体的做法，如果２１天ＴＥＥＲ只有５００多，那细胞出问题的机会比较大．可能是细胞长的太慢，没有铺满（seed cell的数量？）；或者细胞长的不够好，根本形成不了一块儿地毡．
以下是我们用的Procedure，希望有帮助．
Seeding of 24-well culture insert(day1)
*Pre-incubation of the BD transwell culture system:add 200ul of complete DMEM in each insert and 1ml in each well of 24-well plate.Incubate for 2hours at 37℃.
*Measure TEER in blank inserts before seeding.
*Seed 6x10^4 cells/cm^2 (2x10^4 cells/well) to each insert(in triplicate).
*Incubate the transwell culture system in the 37℃ CO2 incubator.
*Change medium for the transwell culture system every other day contiunously for 21-25days.
Remark:
(1)BD BioCoat (Cat No.354803)
Fibrillar collagen TypeI Insert System 24-Multiwell Insert Plate 1.0 Micron.
(2)MILLPORE
MILLICELL-ERS (Cat No.MERS00001)
补充：我查了实验记录，day1空白的TEER是１００多，day21时大部分的TEER已经达到２０００左右了．
给你做个参考，有问题大家在一起讨论．
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请问这个转运的时候这个胶是必须铺的吗？我们用的是康宁的小室。我们还是第一次做，没什么经验，我一个人在摸索，很多地方都很疑惑。请问可否指教一下

因为不知道你们具体的做法，如果２１天ＴＥＥＲ只有５００多，那细胞出问题的机会比较大．可能是细胞长的太慢，没有铺满（seed cell的数量？）；或者细胞长的不够好，根本形成不了一块儿地毡．
以下是我们用的Procedure，希望有帮助．
Seeding of 24-well culture insert(day1)
*Pre-incubation of the BD transwell culture system:add 200ul of complete DMEM in each insert and 1ml in each well of 24-well plate.Incubate for 2hours at 37℃.
*Measure TEER in blank inserts before seeding.
*Seed 6x10^4 cells/cm^2 (2x10^4 cells/well) to each insert(in triplicate).
*Incubate the transwell culture system in the 37℃ CO2 incubator.
*Change medium for the transwell culture system every other day contiunously for 21-25days.
Remark:
(1)BD BioCoat (Cat No.354803)
Fibrillar collagen TypeI Insert System 24-Multiwell Insert Plate 1.0 Micron.
(2)MILLPORE
MILLICELL-ERS (Cat No.MERS00001)
补充：我查了实验记录，day1空白的TEER是１００多，day21时大部分的TEER已经达到２０００左右了．
给你做个参考，有问题大家在一起讨论．
......

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我用的是12well plate。seed 8×10e4 cell/well。肉眼观察，可见颗粒状物质，镜下观察细胞比较密集，但是TEER value 始终上不去，一直是600-700（2k ohmm），单个insert面积为1.12cm^2,所以TEER value应该约是1200k ohmm/cm^2 不知您的2000是什么单位呢！？