[NHCOLL-L:4606] RE: Tissues specimen in ethanol and DMSO-EDTA

[Dirk Neumann]

Hi Carol,

sorry for my late reply, but I would abstain from removing the tissues 
to DMSO-EDTA for practical reasons. It seems, that the DNA-quality 
extracted from DMSO-tissues is bad (problems with degrading resulting in 
short fragments).

A colleague received fish tissues from Russia which were even dried for 
fixation (pinned on a paper and dried in the sun).
During a major project, we had problems with poor quality tubes 
resulting in 100 % evaporation losses which we recognised too late, so 
several tissues dried up. In both cases it was possible to extract high 
quality from the tissues. And prior to lyses of the tissues for 
DNA-extraction, you need to dry the tissues anyway ...

For this reason, and for the forthcoming changes Andy already mentioned, 
I would abstain from transferring the tissues into DMSO-EDTA, as it will 
be difficult to remove residual DMSO from the tissues again.


All the best

Dirk



Bentley, Andrew Charles schrieb:
>
> Hi Carol
>
> I would not be so quick to change your practices. As you may have seen 
> from my previous post to NHCOLL-L about dangerous goods regulations, 
> DOT has recently incorporated what they call “De minimus” regulations 
> into 49 CFR under chapter 173.4b (attached) which now allows for 
> quantities of dangerous goods (ethanol) under 1ml to be excepted from 
> dangerous good regulations. So, at least domestically, you should have 
> no problem sending tissues in less than ml of fluid. We also recently 
> received a letter of interpretation from DOT and USPS stating that 
> packages of scientific specimens packed as per 173.4 are not subject 
> to the regulations (attached). Unfortunately, the international 
> regulations (IATA) have yet to adopt this regulation and even if they 
> did so it would only be in 2013 that this would be written into their 
> regulations and be applicable.
>
> However, I have been working very closely with IATA, DOT and USPS to 
> institute specific regulations for scientific specimens. This is what 
> I wrote in my previous message:
>
> As an aside, I recently received confirmation from my IATA contacts 
> that we have been successful in our bid to have verbiage inserted into 
> the IATA dangerous good manual (Section 2.7) exempting specimen 
> packages from the regulations if packed according to 2.7. The exact 
> verbiage is yet to be drafted but will be along the lines of:
>
> /“Axxx Museum specimens, such as specimens of mammals, birds, 
> amphibians and reptiles, fishes, insects and other invertebrates 
> containing small quantities of UN 1170, UN 1987 or UN 1219 are not 
> subject to these Instructions provided the following packing and 
> marking requirements are met: /
>
> /Specimens are:/
>
> /1. //Wrapped in paper towel and/or cheesecloth moistened with alcohol 
> or an alcohol solution and then placed in a plastic bag that is 
> heat-sealed. Any free liquid in the bag must not exceed 30 mL; or/
>
> /2. //Placed in vials or other rigid containers with no more than 30 
> mL of alcohol or an alcohol solution;/
>
> a) /The prepared specimens are then placed in a plastic bag that is 
> then heat–sealed;/
>
> b) /The bagged specimens are then placed inside another plastic bag 
> with absorbent material then heat sealed;/
>
> c) /The finished bag is then placed in a strong outer packaging with 
> suitable cushioning material;/
>
> d) /The completed package is marked “scientific research specimens, 
> not restricted special provision Axxx applies”./
>
> /The words “not restricted” and the special provision Axxx must be 
> provided on the air waybill when an air waybill is issued./
>
> This will only come into effect in January 2011 but will ensure that, 
> together with the DOT and USPS letters of interpretation that we have 
> already received (attached), all packages, whether domestic or 
> international, will no longer fall into the category of dangerous 
> goods. It will also allow 30ml of free liquid per RIGID internal 
> package (insects and other fragile specimens can be sent in free 
> liquid) and will also allow 95% ethanol (again, up to 30ml free liquid 
> per RIGID internal package).
>
> This has far reaching implications – no training necessary (you can 
> self train to pack), previously inaccessible countries will now be 
> accessible, international mailing by USPS to select countries (based 
> on the USPS International Mailing Manual), no labeling requirements, 
> 30ml of free liquid per internal package, 95% ethanol acceptable etc. 
> It is my hope that this will ease the burden on the small to medium 
> sized museums as well as the larger institutions both within the US 
> and internationally (as most countries follow IATA with some 
> exceptions). This has been a long and arduous process of negotiation, 
> started with our Dangerous Goods roundtable held in Oklahoma, but the 
> light at the end of the tunnel is now blazing!!!
>
> So, unfortunately these too only come into effect in 2011 but you can 
> still make use of excepted quantity regulations IATA 2.7 (also 
> attached) for sending tissues. There are still some problem countries 
> and for those I have been draining all alcohol from the specimens and 
> sending them “dry” as non-dangerous goods. The researcher then 
> rehydrates them upon receipt. We have yet to have any issues with this 
> method and it seems to work just fine. We can get away with this but 
> birds and mammals have infectious regulations (APHIS) that prohibit this.
>
> In terms of your original question – switching tissues from ethanol to 
> DMSO – I would be leery of this due to the extensive osmotic changes 
> that the tissue would go through (although I have no hard evidence to 
> back it up). Just in general terms, I would be hesitant to subject the 
> tissue to any more “trauma” than it has already gone through – who 
> knows what effect this has on DNA strand length etc.
>
> Hope that helps
>
> Andy
>
> A : A : A :
> }<(((_°>.,.,.,.}<(((_°>.,.,.,.}<(((_°>
> V V V
> Andy Bentley
> Ichthyology Collection Manager/Specify Usability Lead
> University of Kansas
> Natural History Museum & Biodiversity Research Center
> Dyche Hall
> 1345 Jayhawk Boulevard
> Lawrence, KS, 66045-7561
> USA
>
> Tel: (785) 864-3863
> Fax: (785) 864-5335
> Email: ABentley at ku.edu
>
> A : A : A :
> }<(((_°>.,.,.,.}<(((_°>.,.,.,.}<(((_°>
> V V V
>
> ------------------------------------------------------------------------
>
> *From:* owner-nhcoll-l at lists.yale.edu 
> [mailto:owner-nhcoll-l at lists.yale.edu] *On Behalf Of *Carol Spencer
> *Sent:* Wednesday, November 04, 2009 8:15 PM
> *To:* NHCOLL-L at lists.yale.edu
> *Subject:* [NHCOLL-L:4591] Tissues specimen in ethanol and DMSO-EDTA
>
> Hi NH-Coll folks:
>
> Restrictions in shipping ethanol and dangerous goods have led us to 
> change out practice, so that now we are sending most tissue samples in 
> buffer (DMSO-EDTA).
>
> Does anybody know of any problems of transferring tissues originally 
> in 95% ethanol or RNALater into DMSO buffer, in terms of quality of 
> DNA for sequencing?
>
> We are also sending tissues originally in RNALater in DMSO-EDTA.
>
> Any thoughts or experiences would be appreciated.
>
> Cheers,
> Carol Spencer
>
> -- 
> Carol L. Spencer
> Staff Curator of Herpetology & Researcher
> Museum of Vertebrate Zoology
> 3101 Valley Life Sciences Building
> University of California, Berkeley, CA, USA 94720-3160
> atrox10 at gmail.com <mailto:atrox10 at gmail.com>
> atrox at berkeley.edu <mailto:atrox at berkeley.edu>
> TEL: 510-643-5778 /FAX: 510-643-8238
>
> http://www.herpnet.org
> http://mvz.berkeley.edu/
>


-- 
Dirk Neumann

Tel: 089 / 8107-111
Fax: 089 / 8107-300
email: Dirk.Neumann(a)zsm.mwn.de

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---------

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email: Dirk.Neumann(a)zsm.mwn.de

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