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人甲狀腺導管癌細胞

1*10^6

thyroid/medulla

Homo sapiens, human

HIV-1、 HBV、HCV、支原體、細菌、酵母和真菌檢測陰性。

MCM-0457：TT細胞專用培養(yǎng)基

epithelial

1:2-1:4

干冰運輸(2ml凍存管)或活細胞運輸（T25細胞瓶）

Freeze medium: 50% basal medium+40% FBS+10%.DMSOStorage temperature: liquid nitrogen vapor phase

adherent

TT cells continuously produce high levels of calcitonin and CEA.Immunoreactive calcitonin was found to be produced in cell culture at levels of 3900 pg/million cells and 7700 pg/million cells 24 and 72 hours respectively, after a medium change.CEA was found to accumulate to greater than 27 ng/million cells over a 72 hours period.Chromosomal analysis of the cell line and tumors induced in nude mice reveal an aneuploid human karyotype with several marker chromosomes.The initial characterization studies of the TT cell line were conducted using early passage TT cells cultivated in RPMI 1640 medium supplemented with 15% fetal bovine serum and 1mM L-glutamine.It is not known if the neuropeptides reported to be produced by this cell line when it was grown in RPMI 1640 medium are also produced by the cells when they are cultured in Ham's F-12K medium.Chromosomal analysis of the cell line and tumors induced in nude mice reveal an aneuploid human karyotype with several marker chromosomes.

如果細胞密度達80%-90%，即可進行傳代培養(yǎng)。1. 棄去培養(yǎng)上清，用不含鈣、鎂離子的PBS潤洗細胞1-2次。2. 加2ml消化液（0.25%Trypsin-0.53mM EDTA）于培養(yǎng)瓶中，置于37℃培養(yǎng)箱中消化1-2分鐘，然后在顯微鏡下觀察細胞消化情況，若細胞大部分變圓并脫落，迅速拿回操作臺，輕敲幾下培養(yǎng)瓶后加少量培養(yǎng)基終止消化。3. 按6-8ml/瓶補加培養(yǎng)基，輕輕打勻后吸出，在1000RPM條件下離心4分鐘，棄去上清液，補加1-2mL培養(yǎng)液后吹勻。4. 將細胞懸液按1：2到1：5的比例分到新的含8ml培養(yǎng)基的新皿中或者瓶中

將含有1mL細胞懸液的凍存管在37℃水浴中迅速搖晃解凍，加入4mL培養(yǎng)基混合均勻。在1000RPM條件下離心4分鐘，棄去上清液，補加1-2mL培養(yǎng)基后吹勻。然后將所有細胞懸液加入培養(yǎng)瓶中培養(yǎng)過夜（或?qū)⒓毎麘乙杭尤?0cm皿中，加入約8ml培養(yǎng)基，培養(yǎng)過夜）。第二天換液并檢查細胞密度。

：待細胞生長狀態(tài)良好時，可進行細胞凍存。下面T25瓶為例；1．細胞凍存時，棄去培養(yǎng)基后，PBS清洗瓶底1-2次后加入1ml胰酶，細胞變圓脫落后，加入2ml完全培養(yǎng)基終止消化，可使用血球計數(shù)板計數(shù)。2．1000RPM離心5分鐘去掉上清。用血清重懸浮，加DMSO至最終濃度為10%。加入DMSO后迅速混勻，按每1ml的數(shù)量分配到凍存管中，注意凍存管做好標識。本公司按每個凍存管細胞數(shù)目大于1X106個細胞凍存。3．將凍存管置于程序降溫盒中，放入-80度冰箱，至少2個小時以后轉(zhuǎn)入液氮灌儲存。記錄凍存管位置以便下次拿取。