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Hi John,
<div class="">yes, that's fair enough. There is, indeed, no reason to suspect that water molecules wouldn't be subject to the same physics as everything else, and motion does explain why you don't see it in the averages. However, the underlying point I was
trying to make is that for purposes of single particle analysis, the "signal" is the perfect projection of a particle, and anything else is noise. Some component of this noise is subject to CTF oscillations, and some component is not. Some component of the
oscillatory term coming from the ice doesn't change this fact. To properly estimate SSNR one needs to actually measure regions of the image not containing particles. I don't believe it can be properly modeled without doing this, since the image background
in single particle data simply isn't "pure water".</div>
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<div class="">Again, though, it isn't clear that having a more accurate SSNR estimate really helps you to get better structures. Yes, theoretically you should weight your data by SSNR, but the practical reality is that the uncertainty in the SSNR estimate makes
it very difficult to achieve actual improvements using this. What SSNR is useful for is evaluating data quality...</div>
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Steven Ludtke, Ph.D.<br class="">
Professor, Dept. of Biochemistry and Mol. Biol. Those who do</div>
<div class="">Co-Director National Center For Macromolecular Imaging<span class="Apple-tab-span" style="white-space: pre; ">
</span> ARE</div>
<div class="">Baylor College of Medicine The converse<br class="">
<a href="mailto:sludtke@bcm.edu" class="">sludtke@bcm.edu</a> -or- <a href="mailto:stevel@alumni.caltech.edu" class="">stevel@alumni.caltech.edu</a> also applies<br class="">
<a href="http://ncmi.bcm.edu/~stevel" class="">http://ncmi.bcm.edu/~stevel</a><br class="">
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<div class="">On Jun 7, 2017, at 8:43 AM, John Rubinstein <<a href="mailto:john.rubinstein@utoronto.ca" class="">john.rubinstein@utoronto.ca</a>> wrote:</div>
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<font size="2" class=""><b class="">***CAUTION:*** This email is not from a BCM Source. Only click links or open attachments you know are safe.</b></font>
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Hi Steve,
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</div>
<div class="">As Greg McMullan and Richard Henderson showed recently, amorphous ice does give Thon rings. However, ice also exhibits a beam induced ‘Brownian’-like movement so that the power spectrum from the average of DDD frames does not often show Thon rings.</div>
<div class=""><a href="https://urldefense.proofpoint.com/v2/url?u=https-3A__www.ncbi.nlm.nih.gov_pubmed_26103047&d=DwMFaQ&c=ZQs-KZ8oxEw0p81sqgiaRA&r=GWA2IF6nkq8sZMXHpp1Xpg&m=bG1ISczT10r6qYocjTXyjaNx3S8A0R8UQjzs20Qymic&s=GMDYOeoavzqGZQPQDUJSg5nbPI14MXeOxMSSmElkzhg&e=" class="">https://www.ncbi.nlm.nih.gov/pubmed/26103047</a></div>
<div class="">Alexis Rohou modified CTF find in order to allow use of this signal in CTF estimation.</div>
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<div class="">Best regards,</div>
<div class="">John</div>
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<div class="">-- </div>
<div class="">John Rubinstein</div>
<div class="">Molecular Medicine Program</div>
<div class="">The Hospital for Sick Children Research Institute</div>
<div class="">686 Bay Street, Rm. 20-9705</div>
<div class="">Toronto, ON</div>
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<div class="">Tel: (+001) 416-813-7255</div>
<div class="">Fax: (+001) 416-813-5022</div>
<div class=""><a href="https://urldefense.proofpoint.com/v2/url?u=http-3A__www.sickkids.ca_research_rubinstein&d=DwMFaQ&c=ZQs-KZ8oxEw0p81sqgiaRA&r=GWA2IF6nkq8sZMXHpp1Xpg&m=bG1ISczT10r6qYocjTXyjaNx3S8A0R8UQjzs20Qymic&s=eKkgq44j8EITMsFv_BXMzKYPNpRvPQUZ23YbxPP-gow&e=" class="">www.sickkids.ca/research/rubinstein</a></div>
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<div class="">On Jun 7, 2017, at 9:33 AM, Ludtke, Steven J <<a href="mailto:sludtke@bcm.edu" class="">sludtke@bcm.edu</a>> wrote:</div>
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Hi Philip,
<div class="">pure water ice, alas, does not exhibit Thon rings (at least not significantly). Other buffer constituents may. This makes modeling noise a bit tricky. </div>
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<div class="">Many of the methods in the field start with the assumption that the image consists of a true projection of a macromolecule, convolved with the CTF with added uncorrelated Gaussian noise. That is, noise with a flat power spectrum and no correlation
among nearby pixels. Prior to direct detectors and counting, the background noise power spectrum in typical cryoEM images had an exponential decay, and could with a little processing be "flattened" to produce something more or less like uncorrelated Gaussian
noise. The problem with this assumption, even then, is that CryoEM images actually contain a lot of other stuff, and from the perspective of single particle reconstruction, noise isn't just the scattering off the solvent, it's anything present in the image
which is not a perfect projection of the particle. </div>
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<div class="">This is what led to EMAN2's approach of computing the power spectrum of masked out particles, and masked out regions outside particles to compute an accurate estimate of the SNR of the particles within the images. Whether having this really produces
better reconstructions is a separate question, but making this estimate DOES tell you quite a lot about particle data quality and whether you should expect to be able to achieve high resolution from your data.</div>
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<div class="">I'm attaching some slides from about 10 years ago showing how SSNR estimates assuming a pretty flat background can give completely inaccurate estimates of the actual SSNR of the images. Hope they are reasonably self-explanatory. This compares
the approach of considering noise to be anything in the power spectrum without Thon rings to be noise, with the approach of computing the noise from regions of the image not containing particles.</div>
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<div class=""><a href="https://urldefense.proofpoint.com/v2/url?u=https-3A__drive.google.com_file_d_0BxreryX3Fp-2DZUVJtTFlYX0lmbzg_view-3Fusp-3Dsharing&d=DwMFaQ&c=ZQs-KZ8oxEw0p81sqgiaRA&r=GWA2IF6nkq8sZMXHpp1Xpg&m=bG1ISczT10r6qYocjTXyjaNx3S8A0R8UQjzs20Qymic&s=vXuqF6vNwv8lPDzq0U9NsFqf83WiofR7JUZ8rhcguxE&e=" class="">https://drive.google.com/file/d/0BxreryX3Fp-ZUVJtTFlYX0lmbzg/view?usp=sharing</a></div>
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<div style="margin: 0px; font-style: normal; font-variant: normal; font-weight: normal; font-size: 12px; line-height: normal; font-family: Courier; " class="">
Steven Ludtke, Ph.D.</div>
<div style="margin: 0px; font-style: normal; font-variant: normal; font-weight: normal; font-size: 12px; line-height: normal; font-family: Courier; " class="">
Professor, Dept of Biochemistry and Mol. Biol. (<a href="http://www.bcm.edu/biochem" class="">www.bcm.edu/biochem</a>)</div>
<div style="margin: 0px; font-style: normal; font-variant: normal; font-weight: normal; font-size: 12px; line-height: normal; font-family: Courier; " class="">
Co-Director National Center For Macromolecular Imaging <span class="Apple-converted-space"> </span>(<a href="http://ncmi.bcm.edu/" class="">ncmi.bcm.edu</a>)</div>
<div style="margin: 0px; font-style: normal; font-variant: normal; font-weight: normal; font-size: 12px; line-height: normal; font-family: Courier; " class="">
Co-Director CIBR Center <span class="Apple-converted-space"> </span>(<a href="http://www.bcm.edu/research/cibr" class="">www.bcm.edu/research/cibr</a>)</div>
<div style="margin: 0px; font-style: normal; font-variant: normal; font-weight: normal; font-size: 12px; line-height: normal; font-family: Courier; " class="">
Baylor College of Medicine<span class="Apple-converted-space"> </span> <span class="Apple-converted-space"> </span></div>
<div style="margin: 0px; font-style: normal; font-variant: normal; font-weight: normal; font-size: 12px; line-height: normal; font-family: Courier; " class="">
<a href="mailto:sludtke@bcm.edu" class="">sludtke@bcm.edu</a></div>
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