Many cultures obtained from a culture collection, such as ECACC, will arrive frozen and in order to use the cells they must be thawed and put into culture. It is vital to thaw cells correctly in order to maintain the viability of the culture and enable the culture to recover more quickly. Some cryoprotectants, such as DMSO, are toxic above 4 °C. Therefore, it is essential that cultures are thawed quickly and diluted in culture medium to minimize the toxic effects.
Materials
- Cell Culture Media: pre-warmed to the appropriate temperature (refer to the ECACC cell line data sheet for the correct medium and temperature.)
- 70% (v/v) alcohol in sterile water (793213)
- DMSO (D2650)
- Trypan Blue Solution (T8154)
Equipment
- Personal protective equipment (sterile gloves, laboratory coat, safety visor)
- Container to transport frozen ampoules e.g. box of dry ice or liquid nitrogen dewar
- Waterbath set to appropriate temperature
- Microbiological safety cabinet at appropriate containment level
- Incubator
- Pre-labelled flasks
- Inverted phase contrast microscope
- Hemocytometer or automated cell counter like a Scepter™ Cell Counter
- Centrifuge
- Marker Pen
- Pipettes
- Ampoule Rack
- Tissue