HeLa cells splitting protocol

Authors

Heike Stichnoth

Materials

Medium

• 450 ml DMEM (GIBCO 1.0 g/l Glucose; Cat. # 31885-023)
• 50 ml FBS (10%, GIBCO Cat. # 10270-106)
• 1 ml Primocin (Invivogen 50mg/ml Cat. # ant-pm-1 )

...sterile filtered through 0.22 μm Milli Pore Bottle Top Filter

Procedure

• remove medium
• wash with PBS (~ 4 -5 ml)
• wash with 0.25 % Tryp./EDTA Solution (use 3 - 4 ml)
• trypsinize cells with 1 ml 0.25 % Tryp./EDTA sol. ~ 5 min/ 37°C
• cells should be about 90 % confluent, if they are less confluent, use less medium to suspend them
• suspend CCL-2 cells in 7 ml medium (total of 8 ml cell suspension) and Kyoto in 9 ml medium (total of 10 ml cell suspension)
• plate 2 x T-75 flasks provided with 15 ml medium
• on Mondays seed out 2 ml cell suspension per flask, on Thursdays 1 ml cell suspension
• go through 20 passages after thawing

Further details

Protocol provided by Heike Stichnoth.

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