[Nhcoll-l] dna storage tubes for -80?C

[Zimkus, Breda Marie]

Dirk,

In response to you inquiry, the text below was posted on the SPNHC list serve by Bryan Stuart of the North Carolina Museum of Natural regarding his experience. I have started a Discussion thread on the SPNHC wiki Genetic Resources Discussion page (https://urldefense.proofpoint.com/v2/url?u=http-3A__spnhc.biowikifarm.net_wiki_Talk-3AGenetic-5FResources&d=CwIFAg&c=-dg2m7zWuuDZ0MUcV7Sdqw&r=CLFZJ3fvGSmDp7xK1dNZfh6uGV_h-8NVlo3fXNoRNzI&m=3hm5Al0bVOAYAz-SsoAOlxRNk5q8b0_4dMY6CrqV8bo&s=UvPUFaJnkFHlkYAFw13dEbTcIZBUYuJkheZdRvtBOEA&e= ) to have this information in one place.

Best,
Breda
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Breda M. Zimkus, Ph.D.
Museum of Comparative Zoology, Harvard University
26 Oxford Street, Cambridge, MA 02138; Tel: 617-496-4656
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In April 2011, the North Carolina Museum of Natural Sciences purchased 1.8 mL NUNC CryoTube vials (cat. no. 368632) for storing tissue collections in our new ultra-cold (-80C) freezers. This vial, which is internally threaded and has a silicone O-ring at the seal, had been used by me (Bryan Stuart) for many years to store amphibian and reptile tissues in EDTA/DMSO salt saturated tissue buffer or in RNAlater (Qiagen), without problems. However, other units at the museum have tissue collections that were preserved in 95-100% ethanol, and we soon discovered after transferring those collections into these vials that ethanol rapidly evaporates from them (on a scale of weeks).

After lodging a complaint with our sales representative, NUNC in Denmark performed an analysis and determined that the vial's silicone O-ring is vapor permeable and therefore not suitable for use with ethanol. In fact, I was also told that none of NUNC and Nalgene's cryogenic vials are intended to store ethanol, but rather were designed for tissue culturing (I broke the news that there are tens or perhaps hundreds of thousands of cryogenic vials being used in natural history collections in the U.S. for non-tissue culture purposes, a large number of which are holding ethanol).

Our sales representative provided us with three alternative vials to test that are externally threaded and lack a silicone O-ring seal (and so are plastic on plastic). These are the Nalgene Cryoware Cryogenic Vials (cat. no. 5000-0020), NUNC CryoTube vials (cat. no. 375418), and Nalgene Cryogenic Vials (cat. no. 5000-1020). In our experiment, we stored 95% ethanol in the original vial with the O-ring and the three alternate makes without an O-ring at four different temperatures (room temperature, 4C, -20C and -80C) for about six weeks. Indeed, the vial with the O-ring dramatically evaporated ethanol, while the other three vials did not. Interestingly, ethanol evaporated fastest at the warmer temperatures. i.e. ethanol evaporated fastest at room temperature and slowest at -80C (I had anticipated the reverse, that freezers were very desiccated and so would evaporate fastest). We also learned that the amount of torque placed on closing the vials with the O-ring seals, i.e. "over-closing" the vials and therefore distorting the O-ring seal, did not have any appreciable effect on evaporation.

In sum, internally threaded cryogenic vials with a silicone O-ring seal such as the NUNC CryoTube vial (cat. no. 368632) should be avoided for storing tissues in ethanol. Instead, we would recommend other models that are externally threaded without an O-ring seal, such as Nalgene Cryoware Cryogenic Vials (cat. no. 5000-0020) or NUNC CryoTube vials (cat. no. 375418).
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Bryan L. Stuart, Ph.D.
Curator of Herpetology & Director of Molecular Genetics Laboratory
North Carolina Museum of Natural Sciences

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