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Dear Nadav,
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<div class="">Could it be that your cryo-EM grid in the column isn’t cold enough, and the contamination originates from formerly vitreous ice which sublimates and re-crystallizes on the surface of the ice layer? </div>
<div class="">Are your holders sufficiently pre-cooled, and are you sure they aren't touching anything warm on the inside of the microscope?</div>
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<div class="">Henning. </div>
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<div class=""><span class="Apple-style-span" style="color: rgb(102, 102, 153); font-family: 'Stone Sans SC ITC TT'; font-size: 11px; ">Henning Stahlberg, PhD</span></div>
<div class=""><font class="Apple-style-span" color="#666699" face="'Stone Sans SC ITC TT'" size="3"><span class="Apple-style-span" style="font-size: 11px; ">Prof. for Structural Biology, C-CINA, Biozentrum, University Basel</span></font></div>
<div class=""><font class="Apple-style-span" color="#666699" face="'Stone Sans SC ITC TT'" size="3"><span class="Apple-style-span" style="font-size: 11px; ">Mattenstrasse 26 | D-BSSE | WRO-1058 | CH-4058 Basel | Switzerland</span></font></div>
<div class=""><font class="Apple-style-span" color="#666699" face="'Stone Sans SC ITC TT'" size="3"><span class="Apple-style-span" style="font-size: 11px; "><a href="http://c-cina.org" class="">http://c-cina.org</a> | Tel. +41-61-387 32 62</span></font></div>
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<div class="">On Sep 1, 2016, at 11:00, Nadav Elad <<a href="mailto:nadav.elad@weizmann.ac.il" class="">nadav.elad@weizmann.ac.il</a>> wrote:</div>
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Dear all<o:p class=""></o:p></div>
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We are having a serious contamination problem in our cryo preparations and would be grateful for any suggestion. Although this issue has been discussed in the past in this forum, the solutions raised do not seem to work. The attached image shows the contamination
on a bad day. I am referring mainly to the lighter, smaller particles (10-20 nm) which are the most persistent. They seem to be above or below the ice layer and are the first to evaporate under the electron beam. The concentration of contaminants is variable.<span class="Apple-converted-space"> </span><o:p class=""></o:p></div>
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<o:p class=""> </o:p></div>
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We are using a Leica EM GP plunger for freezing, and Tecnai T12 and F20 microscopes with Gatan side entry holders. We have tried to replace the ethane cylinder and are in the process of ordering a third one, cleaning the glow discharger machines, different
cryo holders, different cryo-transferring units and grids of varying sources. There seems to be no leaks in the compustages of the microscopes.<o:p class=""></o:p></div>
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<o:p class=""> </o:p></div>
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Thank you for your help<o:p class=""></o:p></div>
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<o:p class=""> </o:p></div>
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Nadav<o:p class=""></o:p></div>
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<o:p class=""> </o:p></div>
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--<o:p class=""></o:p></div>
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Nadav Elad, Ph.D<span dir="RTL" class=""></span><span dir="RTL" class=""></span><span lang="HE" dir="RTL" style="font-family: Arial, sans-serif;" class=""><span dir="RTL" class=""></span><span dir="RTL" class=""></span>.<br class="">
</span>Electron Microscopy Unit<span lang="HE" dir="RTL" style="font-family: Arial, sans-serif;" class=""><br class="">
</span>Weizmann Institute of Science<o:p class=""></o:p></div>
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Rehovot, Israel<span lang="HE" dir="RTL" style="font-family: Arial, sans-serif;" class=""><br class="">
</span>Tel: +972 (0) 8 934 2115<o:p class=""></o:p></div>
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Mobile: +972 (0) 52 3909 420<o:p class=""></o:p></div>
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Email: <span class="Apple-converted-space"> </span><a href="mailto:nadav.elad@weizmann.ac.il" style="color: purple; text-decoration: underline;" class="">nadav.elad@weizmann.ac.il</a><o:p class=""></o:p></div>
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<span id="cid:09B9E3F5-30AD-47D1-ABC7-9115DB46F20D"><contamination.jpg></span><span style="font-family: Helvetica; font-size: 12px; font-style: normal; font-variant-caps: normal; font-weight: normal; letter-spacing: normal; orphans: auto; text-align: start; text-indent: 0px; text-transform: none; white-space: normal; widows: auto; word-spacing: 0px; -webkit-text-stroke-width: 0px; float: none; display: inline !important;" class="">_______________________________________________</span><br style="font-family: Helvetica; font-size: 12px; font-style: normal; font-variant-caps: normal; font-weight: normal; letter-spacing: normal; orphans: auto; text-align: start; text-indent: 0px; text-transform: none; white-space: normal; widows: auto; word-spacing: 0px; -webkit-text-stroke-width: 0px;" class="">
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mailing list</span><br style="font-family: Helvetica; font-size: 12px; font-style: normal; font-variant-caps: normal; font-weight: normal; letter-spacing: normal; orphans: auto; text-align: start; text-indent: 0px; text-transform: none; white-space: normal; widows: auto; word-spacing: 0px; -webkit-text-stroke-width: 0px;" class="">
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