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細胞培養 RPMI Medium 1640

平時研究用的細胞，有需要冷凍保存。冷凍的過程稱為凍存，把凍上的細胞化開的過程叫細胞復蘇。細胞復蘇是一簡單的試驗操作，但操作中有許多需要注意的事項，在實驗操作中注意細小問題，才能使復蘇后的細胞狀態保持良好。

細胞復蘇實驗準備：

主要器材：一次性滴管、打火機、酒精燈、大鑷子、中鑷子、記號筆、廢液缸、封口膜、剪子。

主要試劑： PBS、培養液、消化酶（胰酶）、75%酒精、雙抗。

PBS配制：NaCL :4g   KCL:0.1g   Na2HPO4 :1.745g   KH2PO4 :0.1g   

三蒸水： 500mL，溶解后高壓滅菌。

培養基配制：5mL雙抗、50mL血清，加入培養基中。

細胞復蘇操作步驟：

1.佩戴眼鏡和手套，從液氮罐中取出安瓿或冷凍管。

2.迅速放入38℃水浴中，并不時搖動，在1分鐘內使其*融化，然后在無菌下取出細胞。

3.在1000r/min速度下離心5～10分鐘，棄去上層液，加入適量培養液后接種于培養瓶中，接種濃度1×109/L，置37℃溫箱靜置培養，次日更換一次培養液，繼續培養，觀察生長情況。若細胞密度較高，及時傳代?；驘o需離心直接將細胞加入瓶中，并加入培養基貼壁培養12～24小時后，充去上清，換入新鮮培養基繼續培養。

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細胞培養 RPMI Medium 1640