RESUSCITATION OF FROZEN CELL LINES

LABORATORY PROCEDURES FOR ANIMAL & HUMAN CELL LINES

REFERENCE NO: AHC/1998/3/2.5/1

TITLE: RESUSCITATION OF FROZEN CELL LINES

INTRODUCTION

Frozen cells should not be stored at -80 C for extended periods of time, but in liquid nitrogen (-196 C).

PROCEDURE

Thaw the ampoule in a waterbath (25-37 C).
Transfer the cells to a 10-15 ml centrifuge tube.
Add 6-10 ml prewarmed medium to the cells, mix gently (some scientists recommend dropwise addition of medium).
Centrifuge the cells at 200 x g for 5 min.
Decant supernatant, resuspend the cells in a defined volume of medium, remove a small sample in order to determine cell number and viability, and centrifuge again.
Decant supernatant and adjust cell suspension to the recommended concentration in culture medium containing all supplements listed on the cell line information sheet.

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