Design of a slow cooling device for cryopreservation of small biological samples

Abstract

BACKGROUND: Slow cooling is a cryopreservation methodology where samples are cooled to its storage temperature at controlled cooling rates. OBJECTIVE: Design, construction and evaluation of a simple and low cost device for slow cooling of small biological samples. MATERIALS AND METHODS: The device was constructed based on Pye?s freezer idea. A Dewar flask filled with liquid nitrogen was used as heat sink and a methanol bath containing the sample was cooled at constant rates using copper bars as heat conductor. RESULTS: Sample temperature may be lowered at controlled cooling rate (ranging from 0.4 ºC/min to 6.0 ºC/min) down to ~-60 ºC, where it could be conserved at lower temperatures. An example involving the cryopreservation of Neuro-2A cell line showed a marked influence of cooling rate over post preservation cell viability with optimal values between 2.6 and 4.6 °C/min. CONCLUSION: The cooling device proved to be a valuable alternative to more expensive systems allowing the assessment of different cooling rates to evaluate the optimal condition for cryopreservation of such samples.