Catalogue No.: MZ-2723

Product Format: a T25 flask

 Organism: Mus musculus (mouse)

Complete Growth Medium: See Propagation

Atmosphere: air, 95%; carbon dioxide (CO2), 5% ，37℃

Application:  Cells and cancer research

Propagation: The base medium for this cell line is formulated Eagle's Minimum Essential Medium with 1% NEAA, or Dulbecco's Modified Eagle's Medium with high glucose, add the following components to the base medium: fetal bovine serum to a final concentration of 10%

NOTE: FOR RESEARCH USE ONLY.

 Subculturing: Protocol:

1. Remove and discard culture medium.

2. Briefly rinse the cell layer with 0.25% (w/v) Trypsin - 0.53 mM EDTA solution to remove all traces of serum which contains trypsin inhibitor.

3. Add 2.0 to 3.0 ml of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed (usually within 5 minutes). Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Cells that are difficult to detach may be placed at 37C to facilitate dispersal.

4. Add 6.0 to 8.0ml of complete growthmedium and aspirate cells by gently pipetting.

5. Add appropriate aliquots of the cell suspension to new culture vessels.

6. Incubate cultures at 37C.

Subcultivation Ratio: A subcultivation ratio of 1:3 to 1:5 is recommended

Medium Renewal: 2 to 3 times per week

Tips：

1.     細胞經過運輸后，部分細胞由于溫度變化及劇烈碰撞死亡破碎形成碎片，是正常現象。貼壁細胞可以消化，懸浮細胞直接混勻收集細胞，900-1000rpm(約150g)離心3min，棄上清。加5ml PBS重懸細胞，再900-1000rpm(約150g)離心3min，用新鮮的完全培養基重懸接種到新的培養瓶。第二次PBS重懸是為了去除碎片，如果平時碎片比較少，傳代時可以省略PBS重懸的步驟；如果碎片很多，建議PBS多洗幾次。

2.    細胞生長不均時，可以將細胞消化吹散后加入新的培養基重新接種或傳代。

3.     細胞生長緩慢時，可以選擇提高血清濃度培養（最高不超過20%），也可以根據細胞生長狀態，選擇傳代細胞到新的培養瓶中繼續培養。

4.     不同細胞貼壁性差異比較大，所以消化時間差別較大，20s-10min均有可能，具體以細胞消化到相互分離但未脫落，并可以輕輕吹下為準，嚴禁消化到細胞完全漂浮。客戶消化過度導致細胞死亡、漂浮、生長緩慢，不提供免費售后服務。

5.     干冰發貨均為兩支，客戶先復蘇一支，若復蘇失敗及時聯系我方并在我方指導下復蘇第二支。若客戶同時復蘇兩支均狀態不佳，我方不提供免費售后服務。

6.     細胞狀態正常時，應盡快凍存細胞保種，凍存后應隨機抽取一支檢測凍存效果。我方不對客戶凍存細胞導致死亡負責，客戶凍存細胞死亡不提供免費售后服務。