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I think there are a few causes for concern regarding this paper:

1. Rejuvenation Research isn't a widely-recognized biology journal. If this is the first time memories have been recovered from a cryopreserved organism, this should be in a top journal as it's great news.

2. The article was accepted on the day it was received[ref 1]. This doesn't inspire confidence in the strength of the associated peer review as such minimal review is unusual in biology. Indeed, the average review time for the journal is about 4 weeks[ref 2] (which itself is a short time).

3. This is pretty concerning - the key protocol used in the paper hasn't been published (see reference 14 in the PDF). While it's described in passing in this paper, there's no way to assess the full protocol.

4. There's a juxtaposition between the authors' new technique and an established protocol for freezing C. elegans. However, the authors only freeze C. elegans for 30 minutes in their protocol whereas they freeze organisms in the established protocol testing group for 2 weeks. There seem to be appropriate controls but this difference in protocol is stated very unclearly.

References

1. http://online.liebertpub.com/doi/10.1089/rej.2014.1636

2. http://www.liebertpub.com/DContent/Files/FastFacts/FastFacts...




> If this is the first time memories have been recovered from a cryopreserved organism, this should be in a top journal as it's great news.

Cryonics related research never gets published in top journals no matter its quality, so this is what you would expect. There is a long-standing war by cryobiologists on anything to do with cryonics - the old canards like the exploding lysosomes o' death may have been refuted but last I heard, they were still explicitly banning from membership any cryonicists.

> 2. The article was accepted on the day it was received[ref 1].

Look up the journal policies and note the dates: it was posted the day it was received, as part of Liebert Instant Online/LION http://www.liebertpub.com/forauthors/rejuvenation-research/1... but that doesn't mean it was reviewed or accepted in a day:

> To enable the release of new scientific findings as quickly as possible, the Journal has a policy of pre-publishing all manuscripts in their unedited format upon acceptance. The papers will have undergone full peer review but will not yet have been copyedited, typeset, or proofread/corrected by the authors. All accepted papers appear online within 72 hours of acceptance as a part of Liebert Instant Online (LION).

Given the editing gap is at least October 2014 - April 2015...

> 3. This is pretty concerning - the key protocol used in the paper hasn't been published

Reference 14 is to the SafeSpeed cryopreservation protocol, not to vitrification in general. They also tested out traditional vitrification. If you distrust SafeSpeed, you can throw out that entire part of the paper and the results are exactly the same: preservation of olfactory memory. (This was covered on pg3, I don't know how you missed it if you could look up the reference number...)

> However, the authors only freeze C. elegans for 30 minutes in their protocol whereas they freeze organisms in the established protocol testing group for 2 weeks.

? The 30 minutes refers to the traditional vitrification itself (this is why it's 'slow') where they're immersed in Ln2, but then they were all stored in some more regular freezers at -80 celsius (much warmer than LN2) for several weeks (pg11-12).


> Cryonics related research never gets published in top journals no matter its quality, so this is what you would expect.

In this case it's Aubrey de Grey's own journal, which is getting just a bit incestuous.

Unreviewed or barely-reviewed papers are common these days (e.g. arXiv); this paper's peer review really just isn't up to the peer review standards expected in biology, but that doesn't mean it isn't an interesting result in any case. I'm very glad someone did in fact do this obvious experiment at last.


Wrong! I don't mean to nit-pick, but why are you posting all this inaccurate information? It took several months for blind review of this paper by highly acknowledged scientists. And Rejuvenation Research is a valued scientific research journal. Please ask people directly who are involved with this work before making wrong statements.


> There is a long-standing war by cryobiologists on anything to do with cryonics - the old canards like the exploding lysosomes o' death may have been refuted but last I heard, they were still explicitly banning from membership any cryonicists

I'm not sure what to say here - politics definitely plays into biology publication but there are multiple well-respected journals to send 'small results', of which I don't believe Rejuvenation Research is one.

> Look up the journal policies and note the dates

I did exactly this. Unless Rejuvenation Research has an unusual application of the terms 'Received' and 'Accepted' (see ref. 1), I'm more inclined to believe the information on the article page than a general journal policy.

Weirdly enough, all the articles in the most recently published edition of Rejuvenation Research (http://online.liebertpub.com/toc/rej/18/2) had at least some gap between receipt and acceptance. I'm even more inclined to think something unusual has started to happen with the current batch of online-before-publication papers. This could, of course, be a bug with the journal's content management system.

> Reference 14 is to the SafeSpeed cryopreservation protocol

Publishing a reference to an non-peer-reviewed protocol is unusual, irrespective of the details. This is even more unusual when the protocol forms the core of the paper.

To be a bit more precise (using details from the paper):

Vitrification and slow-freezing are distinct methodologies for preserving specimen. The authors only use two methodologies for preservation, which they explain in the methods section:

    The cryopreservation methods included worms that were trained and cryopreserved by vitrification and worms that were trained and cryopreserved by slow freezing
> If you distrust SafeSpeed, you can throw out that entire part of the paper and the results are exactly the same: preservation of olfactory memory

I think this is part of why my original comment said we should wait for replication and be skeptical. Trust in science is a difficult thing to gain and an easy thing to lose. The choice of journal and the very-unclear description of the cryopreservation protocols make me a bit more skeptical of the results.

The SafeSpeed protocol seems to be under review at Nature Methods. If it's accepted there, the authors will likely need to supply supporting data which will only bolster the claims they made here.

Further, the slow-freezing result is a fairly cheap one to replicate; we're likely to see many people try it and they'll likely cite Alcor's work.

> The 30 minutes refers to the traditional vitrification itself

I think the authors were intentionally vague in comparing the two preservation methods, especially given how clear the rest of the report is.

First, note that only one vitrification technique was used, SafeSpeed. Additionally, only one slow-freezing technique was used, a standard approach that one of your sibling commenters references in passing.

The SafeSpeed-vitrified animals were stored in liquid nitrogen (LN2) for 30 minutes and then recovered. This is stated (unclearly) in the methods section (pg 8) and alluded to in the discussion.

> 30 minutes refers to the traditional vitrification itself (this is why it's 'slow')

This is counter to what the authors state - they explain that the SafeSpeed method is actually slower than 'slow-freezing'. See page 11.

Conclusion

I think that this will be exciting if SafeSpeed gets published somewhere more reputable. As you alluded to earlier, if there is a physical correlate to benzaldehyde training that isn't damaged upon freezing, we'd definitely expect it to be recovered after freezing.

This paper just isn't compelling enough to say that we've seen this occur definitively.

References

1. http://online.liebertpub.com/doi/10.1089/rej.2014.1636

    'Received' usually indicates receipt of a manuscript;
    'Accepted' usually indicates the end of scientific edits.
    Often, articles are held up (i.e., not published online) to coordinate 
    publishing with other labs/projects within a lab.


> I'm not sure what to say here - politics definitely plays into biology publication

This is well beyond regular scientific politics; the attacks are now older than a lot of people working in the field. I don't know why you would expect random journals to be unaffected (where do you get cryobiologist reviewers who are unaffected or fearless?).

> I'm more inclined to believe the information on the article page than a general journal policy.

The journal policy which explains what the dates mean?

> Weirdly enough, all the articles in the most recently published edition of Rejuvenation Research (http://online.liebertpub.com/toc/rej/18/2) had at least some gap between receipt and acceptance.

So are you even objecting to anything besides a software bug?

> The authors only use two methodologies for preservation, which they explain in the methods section:

I don't understand your objection here. OK, so one protocol is still upcoming. I don't think this is that unusual since I see plenty of references to publications in press in papers I read - econ papers spend years as preprints, I regularly comment on drafts, fast moving areas often do this, work which has been turned into multiple publications does this all the time because different journals move at different rates etc. I don't think this is a big problem but even if it was, they also tested out the standard well-understood procedure you cannot possibly have any objection to, so what's with this 'only' business? That's the method one wants them to test - and they did. And it worked the same for preserving memories. Which is the point.

> Further, the slow-freezing result is a fairly cheap one to replicate; we're likely to see many people try it and they'll likely cite Alcor's work.

That seems unlikely. No one wants to go near cryonics for the political reasons I mentioned and there's no funding for it. I have been saying for at least 5 years now that I really hoped someone would run exactly this experiment with seeing if training survived, and it took that long for any study to be done - despite always being cheap to run. Sometimes being cheap to replicate doesn't matter.

> This is counter to what the authors state - they explain that the SafeSpeed method is actually slower than 'slow-freezing'. See page 11.

I don't see any such claim. I think you are again confusing different stages of storage and revival.

> This paper just isn't compelling enough to say that we've seen this occur definitively.

I find it compelling. None of your arguments goes to the root, and most are tangential: it's not a prominent journal because of the very well known politics involved, the dates are due to quirks of the website or acceptance process, the new protocol is not well-described but seems to work as well as the older methods and the older methods suffice to prove the point, and some confusion about the details doesn't obviate the core claim either. The worms were trained, were frozen, were revived, and remembered. Which of your criticisms defeats this? This isn't some mushy social psychology paper with n=15 and the authors could have tried a thousand different analyses to pull out a p<0.05; the worms either remembered or they didn't, there's not many ways to slice the data.


I don't think we're going to agree on the quality of this paper, which is perfectly fine and a normal part of science.

Two things:

> I don't think this is that unusual since I see plenty of references to publications in press in papers I read - econ papers spend years as preprints

This isn't normal in biology. It's a stance that researchers are trying to change (see http://biorxiv.org/, notably), however.

> I don't see any such claim. I think you are again confusing different stages of storage and revival.

At a molecular level, the two processes may work differently (we don't, of course, have the SafeSpeed protocol). However, the article states:

"During this time, it is necessary to allow 30-45 minutes for vitrification processes, 15-30 minutes for slow freezing processes, and 24 hours for all processes to safely recover and verify the survival of the worms"


This paper was peer reviewed and it took many months for it to be published.




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