Detailed Instructions for Thawing Discovery Cryo Leukopaks (RUO)
Materials Needed for Discovery Cryo Leukopaks (RUO) Thawing:
  • HBSS (w/o calcium or magnesium) + 10% FBS + 0.1 mg/ml DNase (Thawing Buffer)
  • ** We recommend the use of DNase to prevent cell clumping due to DNA leakage from dead cells.**
  • 70% Ethanol
  • Discovery Cryo Leukopak (RUO)
  • Sterile bottle
  • Sterile scissors
  • Disposable pipettes and micropipette tips
Equipment Needed for Discovery Cryo Leukopaks (RUO) Thawing:
  • Biological Safety Cabinet
  • Water Bath at 37°C
  • Serological and Micropipettes
  • Liquid Nitrogen Freezer
  • Automated Cell Counter
Protocol:
1

Warm HBSS + 10% FBS in 37C water bath. Wipe with 70% ethanol and transfer into the
biological safety cabinet.
2

Remove Discovery Cryo Leukopak (RUO) from liquid nitrogen storage. Immediately place in a 37°C water bath and fully submerge. Do not move the leukopak while it is thawing.
3

Once only a small ice crystal remains, remove the leukopak from the water bath, thoroughly clean the outside of the bag with 70% ethanol, and transfer into the biological safety cabinet.
4

Using sterile scissors, cut the port on the leukopak and slowly transfer the cell suspension into an appropriate sterile bottle.
5

Add an equal volume of Thawing Buffer to cell suspension dropwise.
6

Add one-half volume of Thawing Buffer to the original leukopak, thoroughly mix, and transfer to cell suspension.
7

Gently add an additional two volumes of Thawing Buffer to the cell suspension.
8

Thoroughly mix the sample, and take a small sample for cell count and viability.
9

Samples are ready to be centrifuged and downstream analyses.
10

Add DNase to 0.1 mg/ml final concentration.