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<div style="direction: ltr;font-family: Tahoma;color: #000000;font-size: 10pt;">Hi Xiaodi and Sharon-<br>
<br>
Here is the script:<br>
<br>
<a href="https://www.dropbox.com/s/l05l48xcz04wrxk/degold.py?dl=0" target="_blank">https://www.dropbox.com/s/l05l48xcz04wrxk/degold.py?dl=0</a><br>
<br>
It uses the EMAN2 image handling library, scipy, and numpy (hopefully you have these installed), and should run very fast even for fairly large images. Let me know if you have any problems.<br>
<br>
Also, I did not try CTFFIND4 at the time. If it can handle gold-containing images as mentioned, it is almost ceratinly a better option than using this program. It could nevertheless be helpful just for looking at the images without funky contrast.<br>
<br>
Cheers,<br>
-Greg<br>
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<p class="p1">Gregory M. Alushin, Ph.D.<br>
Early Independent Scientist<br>
Laboratory of Macromolecular Interactions<br>
Cell Biology and Physiology Center, NHLBI<br>
Building 50, Room 4140<br>
T: (301)-496-6530<br>
E: alushing@mail.nih.gov</p>
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<div style="direction: ltr;" id="divRpF365093"><font color="#000000" face="Tahoma" size="2"><b>From:</b> Reza Khayat [rkhayat@ccny.cuny.edu]<br>
<b>Sent:</b> Thursday, June 09, 2016 2:54 PM<br>
<b>To:</b> Xiaodi Yu; Alushin, Gregory (NIH/NHLBI) [E]; 3dem@ncmir.ucsd.edu<br>
<b>Subject:</b> RE: [3dem] Image processing from the gold grids<br>
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<p class="MsoNormal"><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#1F497D">Hi,</span></p>
<p class="MsoNormal"><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#1F497D"> </span></p>
<p class="MsoNormal"><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#1F497D">The latest version of CTFFIND 4 can deal with the mentioned CTF problems –at least in my hands it does.
</span></p>
<p class="MsoNormal"><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#1F497D"> </span></p>
<p class="MsoNormal"><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#1F497D">Best wishes,<br>
Reza</span></p>
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<p class="MsoNormal"><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#1F497D">Reza Khayat, PhD</span><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#212121"></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#1F497D">Assistant Professor</span><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#212121"></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#1F497D">Department of Chemistry</span><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#212121"></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#1F497D">City College of New York</span></p>
<p class="MsoNormal"><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#1F497D">85 Saint Nicholas Terrace, CDI 2.318</span><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#212121"></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#1F497D">New York, NY 10031</span><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#212121"></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#1F497D"><a href="redir.aspx?REF=b6dtZ9lgtR881_sFzeRfst9DmijPK9LAXciTGqnX8rufqvWwl5DTCAFodHRwOi8vd3d3LmtoYXlhdGxhYi5vcmcv" target="_blank"><span style="color:#0563C1">http://www.khayatlab.org/</span></a></span><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#212121"></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#1F497D">212-650-6070</span><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#212121"></span></p>
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<p class="MsoNormal"><b><span style="font-size:11.0pt; font-family:"Calibri",sans-serif">From:</span></b><span style="font-size:11.0pt; font-family:"Calibri",sans-serif"> 3dem [mailto:3dem-bounces@ncmir.ucsd.edu]
<b>On Behalf Of </b>Xiaodi Yu<br>
<b>Sent:</b> Thursday, June 09, 2016 2:48 PM<br>
<b>To:</b> Alushin, Gregory (NIH/NHLBI) [E] <gregory.alushin@nih.gov>; 3dem@ncmir.ucsd.edu<br>
<b>Subject:</b> Re: [3dem] Image processing from the gold grids</span></p>
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<p style="background:white"><span style="font-family:"Calibri",sans-serif; color:black">Hi Greg Alushin:</span></p>
<p style="background:white"><span style="font-family:"Calibri",sans-serif; color:black"> </span></p>
<p style="background:white"><span style="font-family:"Calibri",sans-serif; color:black">That would be very useful for us. Do you mind to share the link of the program? I would like to have a test for my images.</span></p>
<p style="background:white"><span style="font-family:"Calibri",sans-serif; color:black"> </span></p>
<p style="background:white"><span style="font-family:"Calibri",sans-serif; color:black">Thanks a lot.</span></p>
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<p style="background:white"><span style="font-family:"Calibri",sans-serif; color:black">Xiaodi</span></p>
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<p class="MsoNormal"><b><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:black">From:</span></b><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:black"> 3dem <<a href="redir.aspx?REF=qxMDWeR37vCrnecPMvGPxga_dNkVzYMi3NkBekujW_CfqvWwl5DTCAFtYWlsdG86M2RlbS1ib3VuY2VzQG5jbWlyLnVjc2QuZWR1" target="_blank">3dem-bounces@ncmir.ucsd.edu</a>>
on behalf of Alushin, Gregory (NIH/NHLBI) [E] <<a href="redir.aspx?REF=tulq-IiDLSWqykA3crOZ6wy5uQ7i5SkB01-hW30R9XCfqvWwl5DTCAFtYWlsdG86Z3JlZ29yeS5hbHVzaGluQG5paC5nb3Y." target="_blank">gregory.alushin@nih.gov</a>><br>
<b>Sent:</b> Thursday, June 9, 2016 11:25:17 AM<br>
<b>To:</b> <a href="redir.aspx?REF=aUc9rmFGiQwHF1qg3KnSWX1Aj0_Z6P5IYwnQFqcgw6yfqvWwl5DTCAFtYWlsdG86M2RlbUBuY21pci51Y3NkLmVkdQ.." target="_blank">
3dem@ncmir.ucsd.edu</a><br>
<b>Subject:</b> Re: [3dem] Image processing from the gold grids</span> </p>
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<p class="MsoNormal" style="background:white"><span style="font-size:9.5pt; font-family:"Arial",sans-serif; color:#222222">Hi Xiaodi-
</span></p>
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<p class="MsoNormal" style="background:white"><span style="font-size:9.5pt; font-family:"Arial",sans-serif; color:#222222">Following up on Bob's comment, if you do have gold occupying a significant portion of your image (e.g. the edge of the hole), it can give
you problems with image normalization and CTF estimation with standard practices (programs like CTFFIND). We once collected a dataset where we had enough images of this type that I wrote a small program to try and salvage them. It finds the mean and stdev
of the pixel values in a central 1024 region of the image (which is assumed to be free of gold), then "removes" the gold by replacing pixels more than a certain stdev threshhold with random noise from a distribution that matches the statistics of the central
1024 region. We were able to get OK CTF estimates from these images and they were good for picking particles. Obviously you would want to actually extract particles from the original, unmodified images.</span></p>
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<p class="MsoNormal" style="background:white"><span style="font-size:9.5pt; font-family:"Arial",sans-serif; color:#222222">If you (or anyone else on this list) would find this useful, I'll put up a download link.</span></p>
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<p class="MsoNormal" style="background:white"><span style="font-size:9.5pt; font-family:"Arial",sans-serif; color:#222222">Good luck!</span></p>
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<p class="MsoNormal" style="background:white"><span style="font-size:9.5pt; font-family:"Arial",sans-serif; color:#222222">-Greg Alushin</span></p>
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<p class="MsoNormal" style="background:white"><span style="font-size:9.5pt; font-family:"Arial",sans-serif; color:#222222">On Thu, Jun 9, 2016 at 9:08 AM, Grassucci, Robert <<a href="redir.aspx?REF=n5py8DcuEBdcfp7VcsyRBMPB4rWNWQaFlPQn-yhvBC2fqvWwl5DTCAFodHRwczovL21haWwubmloLmdvdi9vd2EvcmVkaXIuYXNweD9SRUY9RTd3U0RtQUwwWVRWdkVYUmJldktIRFQ4U0FGRGh6QzZ3YXZuX0ozd083dUt0aUJOazVEVENBRnRZV2xzZEc4NmNtY3lOVEF5UUdOMWJXTXVZMjlzZFcxaWFXRXVaV1Ix" target="_blank"><span style="color:#1155CC">rg2502@cumc.columbia.edu</span></a>> wrote:</span></p>
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<p class="MsoNormal" style="background:white"><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#1F497D">Dear Xiaodi,</span><span style="color:#222222"></span></p>
<p class="MsoNormal" style="background:white"><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#1F497D">What you may be seeing is normalization/display issue if the gold is a significant portion of your image. Of course I am assuming
that your ice is the same thickness in both types of grids. In our hands the holey gold grids do seem to require a slightly longer blot time so that could also be the problem. We have also noticed some gold particles on both the commercial and home-made
holey gold grids but as long as it does not significantly alter the overall standard deviation it is not a problem . I hope that helps you.</span><span style="color:#222222"></span></p>
<p class="MsoNormal" style="background:white"><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#1F497D">Best,</span><span style="color:#222222"></span></p>
<p class="MsoNormal" style="background:white"><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#1F497D">Bob</span><span style="color:#222222"></span></p>
<p class="MsoNormal" style="background:white"><span style="font-size:11.0pt; font-family:"Calibri",sans-serif; color:#1F497D"> </span><span style="color:#222222"></span></p>
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<p class="MsoNormal" style="background:white"><b><span style="font-size:10.0pt; font-family:"Tahoma",sans-serif; color:#222222">From:</span></b><span style="font-size:10.0pt; font-family:"Tahoma",sans-serif; color:#222222"> 3dem [mailto:<a href="redir.aspx?REF=_VuYGXWEMcmJKqkCDvIa4pZT0gqexJ6IuBZntKgBOH2fqvWwl5DTCAFodHRwczovL21haWwubmloLmdvdi9vd2EvcmVkaXIuYXNweD9SRUY9bXh4S3ltemVZSk95V2pXbzFJekhnSFBLdDVESE5vYWFhNzJxVjNWZ1VTQ0t0aUJOazVEVENBRnRZV2xzZEc4Nk0yUmxiUzFpYjNWdVkyVnpRRzVqYldseUxuVmpjMlF1WldSMQ.." target="_blank"><span style="color:#1155CC">3dem-bounces@ncmir.ucsd.edu</span></a>] <b>On
Behalf Of </b>Xiaodi Yu<br>
<b>Sent:</b> Wednesday, June 08, 2016 11:27 PM<br>
<b>To:</b> <a href="redir.aspx?REF=0zLvTdHVHEFc5yi_200dDt4424V2xskr_vPLnpw3XcKfqvWwl5DTCAFodHRwczovL21haWwubmloLmdvdi9vd2EvcmVkaXIuYXNweD9SRUY9WVM0aXZWQ1RUV0Rxd3E3ZDEyV0tqUV82blRTcHQzRWh0eGMzd3ZBTXNXR0t0aUJOazVEVENBRnRZV2xzZEc4Nk0yUmxiVUJ1WTIxcGNpNTFZM05rTG1Wa2RRLi4." target="_blank"><span style="color:#1155CC">3dem@ncmir.ucsd.edu</span></a><br>
<b>Subject:</b> [3dem] Image processing from the gold grids</span><span style="color:#222222"></span></p>
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<p style="background:white"><span style="font-size:9.5pt; font-family:"Calibri",sans-serif; color:black"> </span><span style="font-size:9.5pt; font-family:"Arial",sans-serif; color:#222222"></span></p>
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<p style="background:white"><span style="font-size:9.5pt; font-family:"Calibri",sans-serif; color:black">Dear All:</span><span style="font-size:9.5pt; font-family:"Arial",sans-serif; color:#222222"></span></p>
<p style="background:white"><span style="font-size:9.5pt; font-family:"Calibri",sans-serif; color:black"> </span><span style="font-size:9.5pt; font-family:"Arial",sans-serif; color:#222222"></span></p>
<p style="background:white"><span style="font-size:9.5pt; font-family:"Calibri",sans-serif; color:black">We have one dataset collected from the gold grids. We feel that the particle contrast from the gold grids is weaker comparing with the one from the regular
carbon grids. We do find some gold particles in the images. So we wonder is it possible that these gold particles affect the image normalization/contrast? And is any prefilter or re-adjust the normalization of the images needed before we process the dataset?
The last question is comparing with the carbon grids, do the gold grids prefer longer blotting time in order to get the thin ice?</span><span style="font-size:9.5pt; font-family:"Arial",sans-serif; color:#222222"></span></p>
<p style="background:white"><span style="font-size:9.5pt; font-family:"Calibri",sans-serif; color:black"> </span><span style="font-size:9.5pt; font-family:"Arial",sans-serif; color:#222222"></span></p>
<p style="background:white"><span style="font-size:9.5pt; font-family:"Calibri",sans-serif; color:black">This is the first time we collected the data from the gold grids. So any suggestion is welcome. Thank you in advanced.</span><span style="font-size:9.5pt; font-family:"Arial",sans-serif; color:#222222"></span></p>
<p style="background:white"><span style="font-size:9.5pt; font-family:"Calibri",sans-serif; color:black"> </span><span style="font-size:9.5pt; font-family:"Arial",sans-serif; color:#222222"></span></p>
<p style="background:white"><span style="font-size:9.5pt; font-family:"Calibri",sans-serif; color:black">Best ,</span><span style="font-size:9.5pt; font-family:"Arial",sans-serif; color:#222222"></span></p>
<p style="background:white"><span style="font-size:9.5pt; font-family:"Calibri",sans-serif; color:black"> </span><span style="font-size:9.5pt; font-family:"Arial",sans-serif; color:#222222"></span></p>
<p style="background:white"><span style="font-size:9.5pt; font-family:"Calibri",sans-serif; color:black">Xiaodi</span><span style="font-size:9.5pt; font-family:"Arial",sans-serif; color:#222222"></span></p>
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<p class="p1"><span style="font-size:10.0pt; font-family:"Tahoma",sans-serif; color:black">Gregory M. Alushin, Ph.D.<br>
Early Independent Scientist<br>
Laboratory of Macromolecular Interactions<br>
Cell Biology and Physiology Center, NHLBI<br>
Building 50, Room 4140<br>
T: (301)-496-6530<br>
E: <a href="redir.aspx?REF=QA3OJDW4AM-5Y3jK981bfH_0T2KXWrY7eb3xRKYxMJifqvWwl5DTCAFtYWlsdG86YWx1c2hpbmdAbWFpbC5uaWguZ292" target="_blank">
alushing@mail.nih.gov</a></span></p>
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