<div dir="ltr"><div class="gmail_default" style="font-family:trebuchet ms,sans-serif;font-size:small">Hello all,</div><div class="gmail_default" style="font-family:trebuchet ms,sans-serif;font-size:small"><br></div><div class="gmail_default" style="font-family:trebuchet ms,sans-serif;font-size:small">Is it possible it is a disordered sponge phase that formed from micelles during freezing?</div><div class="gmail_default" style="font-family:trebuchet ms,sans-serif;font-size:small"><br></div><div class="gmail_default" style="font-family:trebuchet ms,sans-serif;font-size:small">Best, Howard</div></div><br><div class="gmail_quote"><div dir="ltr" class="gmail_attr">On Thu, Apr 13, 2023 at 8:37 AM Mariena Silvestry Ramos <<a href="mailto:ms3289@cornell.edu">ms3289@cornell.edu</a>> wrote:<br></div><blockquote class="gmail_quote" style="margin:0px 0px 0px 0.8ex;border-left:1px solid rgb(204,204,204);padding-left:1ex">
<div dir="auto">
Hi James and all,
<div><br>
</div>
<div>I agree. I've seen "webby" stuff like that too, even in protein preps, which I've attributed in the past to denatured/mesbehaving protein. Unfortunately I don't have an x-ray detector on any of my columns to quickly check what's in them, but that would
be a possibility in a system that does have it. </div>
<div><br>
</div>
<div>The items in question do look like micelles, and as Vincent pointed out, we don't have info on what the contents/combo of lipids and detergents are. </div>
<div><br>
</div>
<div>Best,</div>
<div><br>
</div>
<div>Mariena<br>
<br>
<div dir="ltr">Sent from my Game Boy</div>
<div dir="ltr"><br>
<blockquote type="cite">On Apr 13, 2023, at 10:08 AM, David Michael Belnap <<a href="mailto:David.Belnap@utah.edu" target="_blank">David.Belnap@utah.edu</a>> wrote:<br>
<br>
</blockquote>
</div>
<blockquote type="cite">
<div dir="ltr">
<div>
<p class="MsoNormal"><span style="font-family:Georgia,serif">James,<u></u><u></u></span></p>
<p class="MsoNormal"><span style="font-family:Georgia,serif"><u></u> <u></u></span></p>
<p class="MsoNormal"><span style="font-family:Georgia,serif">Those images look consistent with large particles that exclude water. I don’t know the expected sizes of micelles, however. If the appearance changes with increased exposure to the electron beam,
then I would agree that you have beam damage. Be careful of claims of “pure” samples. I often see much more than what is claimed to be in a pure sample. Is what you are looking for supposed to be smaller than these particles? As Vincent suggests, you may
have constructs different from what you expect. Your lower magnification picture suggests a diluted sample to me.<u></u><u></u></span></p>
<p class="MsoNormal"><span style="font-family:Georgia,serif"><u></u> <u></u></span></p>
<p class="MsoNormal"><span style="font-family:Georgia,serif">David<u></u><u></u></span></p>
<p class="MsoNormal"><span style="font-family:Georgia,serif"><u></u> <u></u></span></p>
<p class="MsoNormal"><span style="font-family:Georgia,serif"><u></u> <u></u></span></p>
<div style="border-right:none;border-bottom:none;border-left:none;border-top:1pt solid rgb(181,196,223);padding:3pt 0in 0in">
<p class="MsoNormal"><b><span style="font-size:12pt;color:black">From: </span></b><span style="font-size:12pt;color:black">3dem <<a href="mailto:3dem-bounces@ncmir.ucsd.edu" target="_blank">3dem-bounces@ncmir.ucsd.edu</a>> on behalf of vincent Chaptal <<a href="mailto:vincent.chaptal@ibcp.fr" target="_blank">vincent.chaptal@ibcp.fr</a>><br>
<b>Date: </b>Thursday, April 13, 2023 at 7:58 AM<br>
<b>To: </b>"Kizziah, James" <<a href="mailto:kizziah4@uab.edu" target="_blank">kizziah4@uab.edu</a>>, "<a href="mailto:3dem@ncmir.ucsd.edu" target="_blank">3dem@ncmir.ucsd.edu</a>" <<a href="mailto:3dem@ncmir.ucsd.edu" target="_blank">3dem@ncmir.ucsd.edu</a>><br>
<b>Subject: </b>Re: [3dem] Interpretation of strange objects in a cryo-EM sample<u></u><u></u></span></p>
</div>
<div>
<p class="MsoNormal"><u></u> <u></u></p>
</div>
<p class="MsoNormal" style="margin-bottom:12pt"><span style="font-size:13.5pt;font-family:"Courier New"">Dear James,
<br>
<br>
you didn't say which detergent the micelles would be orinigating from, it changes from detergent to detergent.
<br>
<br>
But in any case, beside the mix DDM/CHS which forms small micelles when concentrated but large objects when diluted, most micelles are much much smaller, say 30-50Å or so.
<br>
<br>
I think there is an article by the group of Arne Moeller on detergent micelle observation by negative stain, I don't recall if they tried in cryo.
<br>
<br>
Best<br>
Vincent</span><u></u><u></u></p>
<div>
<p class="MsoNormal">Le 13/04/2023 à 15:48, Kizziah, James a écrit :<u></u><u></u></p>
</div>
<blockquote style="margin-top:5pt;margin-bottom:5pt">
<div>
<p class="MsoNormal" style="line-height:120%">
Hi all,<u></u><u></u></p>
<p class="MsoNormal" style="line-height:120%">
<u></u><u></u></p>
<p class="MsoNormal" style="line-height:120%">
We have imaged a cryo-EM sample presumed to contain micelles and observed the large, white objects full of/covered with junk shown in the attached images. Our best guess is that these are (or were) the micelles and have been destroyed by the electron beam,
contact with the air water interface, or both, after vitrification. Has anyone seen something like this before or does anyone have alternative interpretations?
<u></u><u></u></p>
<p class="MsoNormal" style="line-height:120%">
<u></u><u></u></p>
<p class="MsoNormal" style="line-height:120%">
Glow-discharged lacey carbon grids plunge frozen with a Mark IV Vitrobot, clipped and imaged on a Glacios.
<u></u><u></u></p>
<p class="MsoNormal" style="line-height:120%">
<u></u><u></u></p>
<p class="MsoNormal" style="line-height:120%">
Thanks,<u></u><u></u></p>
<p class="MsoNormal" style="line-height:120%">
James<u></u><u></u></p>
</div>
<p class="MsoNormal"><br>
<br>
<u></u><u></u></p>
<pre>_______________________________________________<u></u><u></u></pre>
<pre>3dem mailing list<u></u><u></u></pre>
<pre><a href="mailto:3dem@ncmir.ucsd.edu" target="_blank">3dem@ncmir.ucsd.edu</a><u></u><u></u></pre>
<pre><a href="https://mail.ncmir.ucsd.edu/mailman/listinfo/3dem" target="_blank">https://mail.ncmir.ucsd.edu/mailman/listinfo/3dem</a><u></u><u></u></pre>
</blockquote>
<p class="MsoNormal"><u></u> <u></u></p>
<div>
<p class="MsoNormal">-- <br>
<br>
<u></u><u></u></p>
<p><span>Vincent Chaptal, PhD</span><u></u><u></u></p>
<p><span>Director of GdR APPICOM</span><u></u><u></u></p>
<p><span>Drug Resistance and Membrane Proteins Lab</span><u></u><u></u></p>
<p><u></u> <u></u></p>
<p><span>MMSB -UMR5086</span><u></u><u></u></p>
<p><span>7 passage du Vercors</span><span> </span><u></u><u></u></p>
<p><span>69007 LYON</span><u></u><u></u></p>
<p><span>FRANCE</span><u></u><u></u></p>
<p><span>+33 4 37 65 29 01</span><u></u><u></u></p>
<p><span><a href="https://urldefense.com/v3/__http:/www.appicom.cnrs.fr__;!!Mih3wA!A3tyGeCKHy1u_ntKsI6Sj44WCXHLXr90Zthw3sl9RHWJ31CgkVaeoByZhSXzEiLqXUj5So3w4Nl7c4Gbthk2_aHTE3EvKQ$" target="_blank">http://www.appicom.cnrs.fr</a></span><u></u><u></u></p>
<p><span><a href="https://urldefense.com/v3/__http:/mmsb.cnrs.fr/en/__;!!Mih3wA!A3tyGeCKHy1u_ntKsI6Sj44WCXHLXr90Zthw3sl9RHWJ31CgkVaeoByZhSXzEiLqXUj5So3w4Nl7c4Gbthk2_aHYmG3vpQ$" target="_blank">http://mmsb.cnrs.fr/en/</a></span><u></u><u></u></p>
<p><u></u> <u></u></p>
</div>
</div>
<span>_______________________________________________</span><br>
<span>3dem mailing list</span><br>
<span><a href="mailto:3dem@ncmir.ucsd.edu" target="_blank">3dem@ncmir.ucsd.edu</a></span><br>
<span><a href="https://mail.ncmir.ucsd.edu/mailman/listinfo/3dem" target="_blank">https://mail.ncmir.ucsd.edu/mailman/listinfo/3dem</a></span><br>
</div>
</blockquote>
</div>
</div>
_______________________________________________<br>
3dem mailing list<br>
<a href="mailto:3dem@ncmir.ucsd.edu" target="_blank">3dem@ncmir.ucsd.edu</a><br>
<a href="https://mail.ncmir.ucsd.edu/mailman/listinfo/3dem" rel="noreferrer" target="_blank">https://mail.ncmir.ucsd.edu/mailman/listinfo/3dem</a><br>
</blockquote></div><br clear="all"><div><br></div><span class="gmail_signature_prefix">-- </span><br><div dir="ltr" class="gmail_signature"><div dir="ltr"><div dir="ltr"><div dir="ltr"><div dir="ltr"><div dir="ltr"><div dir="ltr"><div dir="ltr"><div dir="ltr"><div dir="ltr"><div dir="ltr"><div dir="ltr"><div dir="ltr"><div dir="ltr"><div dir="ltr"><font size="2"><span style="font-family:garamond,serif">Howard S. Young, PhD</span><br style="font-family:garamond,serif"><span style="font-family:garamond,serif">Professor</span><br style="font-family:garamond,serif"><span style="font-family:garamond,serif">Biochemistry, MSB 3-27<br>University of Alberta</span><br style="font-family:garamond,serif"><span style="font-family:garamond,serif">Edmonton, Alberta T6G 2H7</span><br></font></div><div dir="ltr"><font size="2"><span style="font-family:garamond,serif"><br></span></font></div><div><span style="font-family:garamond,serif"><b><a href="https://urldefense.com/v3/__https://sites.google.com/ualberta.ca/mpdrg/home__;!!Mih3wA!HupVjI7D3RNV1qC9hVduHwrUSD_oWmHSntEXLvTqfxuYruUMZ1n_tzGS78i_AZ2YSNRqyix_I58S-Kil8Xn9$" target="_blank"><font size="4">Membrane Protein Disease Research Group</font></a></b></span></div></div></div></div></div></div></div></div></div></div></div></div></div></div></div>