| 產(chǎn)品名稱 |
NIT-1 |
| 商品貨號 |
MZ-0288 |
| 中文名稱 |
小鼠胰島素瘤β細胞 |
| 細胞數(shù)量 |
1*10^6 |
| 組織來源 |
pancreas/islet of langerhans |
| 細胞種屬 |
mus musculus, transgenic for sv40 large t antigen, mouse, transgenic for sv40 large t antigen |
| 細胞污染 |
HIV-1、 HBV、HCV、支原體、細菌、酵母和真菌檢測陰性。 |
| 生長特性 |
adherent |
| 培養(yǎng)基 |
Ham's F-12K+10% heat-inactivated dialyzed FBS+1% P/S |
| 形態(tài)特征 |
epithelial |
| 傳代方法 |
1:2-1:3 |
| 培養(yǎng)條件 |
Atmosphere: Air, 95%; CO2, 5%。Temperature: 37℃ |
| 細胞描述 |
These mice are transgenic for the SV40 large T antigen under the control of a rat Insulin promoter, and spontaneously develop beta adenomas.At passage 18, most cells stained positively for Insulin, less than 5% were positive for glucagon and none were positive for somatostatin or pancreatic polypeptide.Insulin secretion is responsive to glucose concentration in the medium.There is low constitutive expression of MHC class I, class Ⅱ and ICAM-1 mRNA, but expression of both is markedly increased by treatment with interferon gamma.Stimulation of class I mRNA is accompanied by increased class I antigen expression and induction of an occult class I product expressing the H-2.39 specificity.MHC class Ⅱ antigen is not induced despite the induction of the mRNA.NIT-1 cells show ultrastructural features of differentiated mouse beta cells(well developed rough endoplasmic reticulum, extensive golgi apparatus and beta granules).The cells shed a mature ecotropic type C retrovirus. The retrovirus is capable of infecting other Fv-1 n mouse cell lines, so care should be taken to avoid cross infection. NOTE: NIT-1 cells will not form a confluent monolayer; however, they will form nice colonies of monolayered cells in a fairly dense array.When the NIT-1 colonies begin to ball up slightly and show many round cells on top of the monolayers as well as floating in the media, it is time to passage them. |
| 細胞傳代步驟 |
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 |
| 復(fù)蘇細胞步驟 |
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。 |
| 細胞凍存步驟 |
：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。 |
| 細胞凍存 |
Freeze medium: 50% basal medium+40% FBS+10%.DMSOStorage temperature: liquid nitrogen vapor phase |
| 細胞運輸 |
干冰運輸(2ml凍存管)或活細胞運輸（T25細胞瓶） |
