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<p>If you would please advise on the following:<br>
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<p>I have protein that shows strong preferential orientation. The buffer only contains (20 mM Tris. HCl pH7.5, 150 mM NaCl). </p>
<p>I tried adding detergent CHAPS (8 mM final concentration) and the number of particles dropped to only a few per hole and most of the protein particels were on the carbon film. <span style="font-family:Calibri,Arial,Helvetica,sans-serif; font-size:16px; background-color:rgb(255,255,255)">I
tried different hole size gold grids r</span><span style="font-family:Calibri,Arial,Helvetica,sans-serif; font-size:16px; background-color:rgb(255,255,255)">1.2/1.3</span><span style="font-family:Calibri,Arial,Helvetica,sans-serif; font-size:16px; background-color:rgb(255,255,255)"> and
r</span><span style="font-family:Calibri,Arial,Helvetica,sans-serif; font-size:16px; background-color:rgb(255,255,255)">2/2 but the same result. Any advice how to obtain random distribution in such case.</span></p>
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<p>Is there an adjustment in Relion 3d classification parameters in order to obtain a 3d classification? I tried without symmetry and both the initial model and 3d classifcation fail. <br>
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<p><span style="font-size: 12pt;">number of classes</span><br>
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<p><span style="font-size:11.5pt; line-height:107%; font-family:"Segoe UI",sans-serif; color:#212121">Initial angular sampling</span><br>
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<p>symmetry<br>
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<p>CTF correction or 'Ignore CTF’s until first peak'.</p>
<p><span style="font-size: 12pt;">'Initial angular sampling'.</span></p>
<p>mask<br>
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<p>SGD:<br>
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<p>Final resolution<br>
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<p>Initial mini-batch size<br>
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<p><span style="font-size:11.5pt; line-height:107%; font-family:"Segoe UI",sans-serif; color:#212121"><br>
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<p><span style="font-family:Calibri,Arial,Helvetica,sans-serif; font-size:16px; background-color:rgb(255,255,255)">Thank you very much for your help.</span><br>
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