[3dem] [ccpem] Adequate particle normalization prior to 2D classification?
Edward Egelman
egelman at virginia.edu
Sun Jan 31 13:20:45 PST 2016
I would not have corrected you, just Marin!
Regards,
Ed
On 1/31/16 4:18 PM, Nancy Meyer wrote:
> Well, since *I'm* the guilty "people" (and apparently have company), I
> thought I should say that it was a casual error and that I'd no reason
> to think Schors wouldn't know exactly what I meant. Apologies for my
> unfamiliarity mostly to Scientific Reports. Glad we're keeping
> pretentiousness in check. And that ya'll are having this debate on
> Sunday. ;-) But thanks in all sincerity for the dialogue - I stand
> corrected!
>
> Very best,
> Nancy
>
> Sent from my iPhone
>
> On Jan 31, 2016, at 8:35 AM, Edward Egelman <egelman at virginia.edu
> <mailto:egelman at virginia.edu>> wrote:
>
>> Why are people using "Nature, Scientific Reports"? The journal's
>> name is Scientific Reports, and Nature is the publisher. By this
>> logic I could say that I was the Editor-in-Chief of Cell, Biophysical
>> Journal, since Cell is the publisher of BJ. Not to rain on Marin's
>> parade, but If one looks at Peer Review Policy for Nature
>> publications, it states:
>> "The policy outlined on this page applies to Nature journals (those
>> with the word "Nature" in their title). NPG publishes many other
>> journals, each of which has separate publication policies described
>> on its website."
>> The policy for Nature journals goes on to explain the need for
>> novelty, significance and likely impact. In stark contrast, this is
>> what they say about Scientific Reports:
>> "To be considered for publication in Scientific Reports, a paper must
>> be technically sound original research, without any requirement for
>> impact or a conceptual advance."
>> Just my two cents (which are worth less and less).
>> Ed
>>
>> On 1/30/16 8:07 AM, Marin van Heel wrote:
>>>
>>> Dear Nancy,
>>>
>>> What Sjors is suggesting is a simplified version of what we
>>> suggested in:
>>>
>>> "A posteriori correction of camera characteristics from large image
>>> data sets", P. Afanasyev et al, Nature, Scientific Reports.
>>>
>>> The camera correction should preferably include the subtraction of
>>> the average background and the normalization of the standard
>>> deviations of the pixel vectors, especially if you are operating
>>> your DE camera in integrating mode. The movie alignments can improve
>>> significantly after the a posteriori camera correction.The other
>>> aspect is the FRC validation of the camera correction. The FRC is
>>> then not used as a ‘gold standard’ 2D resolution criterion but
>>> rather as an indicator of the independence of different images
>>> collected with the same sensor. The FRC should not cross the 3-sigma
>>> threshold indicating the expected random noise fluctuations around
>>> FRC=0. For details see the paper.
>>>
>>> If you want to test the a posteriori camera correction programs,
>>> contact michael at imagescience.de!
>>>
>>> Hope this helps!
>>>
>>> Marin
>>>
>>> =====================================
>>>
>>> On 28/01/2016 00:48, Nancy Meyer wrote:
>>>> Again, thank you for the reassuring response! Maybe I've lucked out with similar image intensities in previous datasets - I should probably look at a few min/max, mean, and stdev values to convince myself.
>>>>
>>>> I was pretty careful to remove particles on that horiz artefact and if restoring their inclusion is the biggest gain from redoing gain correction, I can maybe let that slide for now. I did just see "A posteriori correction of camera
>>>> characteristics from large image data sets", P. Afanasyev et al, Nature, Scientific Reports, after looking up your recommendation - looks like they've implemented it in Imagic - wonder if you could do the same correction in just ImageJ...
>>>>
>>>> Thank you for taking the time to look at the images... slowly getting the hang of Relion, but the input is so invaluable!
>>>>
>>>> Best,
>>>> Nancy
>>>>
>>>> ______________________________________
>>>> From: Sjors Scheres [scheres at mrc-lmb.cam.ac.uk]
>>>> Sent: Wednesday, January 27, 2016 1:10 PM
>>>> To: Nancy Meyer
>>>> Cc:ccpem at jiscmail.ac.uk
>>>> Subject: Re: [ccpem] Adequate particle normalization prior to 2D classification?
>>>>
>>>> Hi Nancy,
>>>> The particles look perfectly fine to me. The overall differences in
>>>> greyscale you see is because each of the particles is scaled from white to
>>>> black with their own min and max values, which are susceptible to
>>>> outliers. The mean will probably be perfectly fine. You can get rid of
>>>> this with removing white and black dust, but it will not be necessary to
>>>> do so.
>>>>
>>>> The classes also look quite OK. Just select all the nice ones and proceed
>>>> with another 2D classification or go into 3D already. Some details are
>>>> already coming up, so the 3D structure will look nice.
>>>>
>>>> The micrographs show a sign of a bad gain correction. If you have many of
>>>> them (> several hundreds) you could correct the gain a-posteriori by
>>>> calculating the average of all micrographs and then divide each micrograph
>>>> by that average. This should get rid of the horizontal artefact (as long
>>>> as it is present in all micrographs). That may save some particles that
>>>> lie on that line.
>>>>
>>>> HTH,
>>>> Sjors
>>>>
>>>>
>>>>
>>>>
>>>
>>> --
>>> ================================================================
>>>
>>> Prof Dr Ir Marin van Heel
>>>
>>> Professor of Cryo-EM Data Processing
>>>
>>> Leiden University
>>> NeCEN Building Room 05.27
>>> Einsteinweg 55
>>> 2333 CC Leiden
>>> The Netherlands
>>>
>>> Tel. NL: +31(0)715271424 // Mobile NL: +31(0)652736618
>>> Skype: Marin.van.Heel
>>> email: marin.vanheel(A_T)gmail.com <http://gmail.com>
>>> and: mvh.office(A_T)gmail.com <http://gmail.com>
>>>
>>>
>>>
>>> _______________________________________________
>>> 3dem mailing list
>>> 3dem at ncmir.ucsd.edu
>>> https://mail.ncmir.ucsd.edu/mailman/listinfo/3dem
>>
>> --
>>
>>
>>
>> Edward H. Egelman, Ph.D.
>>
>> Harrison Distinguished Professor
>>
>> Dept. of Biochemistry and Molecular Genetics
>>
>> University of Virginia
>>
>> President
>>
>> Biophysical Society
>>
>> phone: 434-924-8210
>>
>> fax: 434-924-5069
>>
>> egelman at virginia.edu
>>
>> http://www.people.virginia.edu/~ehe2n
>>
>>
>> _______________________________________________
>> 3dem mailing list
>> 3dem at ncmir.ucsd.edu <mailto:3dem at ncmir.ucsd.edu>
>> https://mail.ncmir.ucsd.edu/mailman/listinfo/3dem
--
Edward H. Egelman, Ph.D.
Harrison Distinguished Professor
Dept. of Biochemistry and Molecular Genetics
University of Virginia
President
Biophysical Society
phone: 434-924-8210
fax: 434-924-5069
egelman at virginia.edu
http://www.people.virginia.edu/~ehe2n
<http://www.people.virginia.edu/%7Eehe2n>
icture
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