<div dir="ltr"><div>Hi Sylvain,</div><div><br></div><div>>
Or maybe this is a flip and then the orientation would be ok, but this would change the handedness. <br></div><div>Yes, this is what I think might be happening here. This "off by just 10 degrees" is misleading, it's actually the expected 5 degrees deviation from vertical, but going over to the other side of the axis. Your tilt series images are probably being flipped somewhere in your pipeline (perhaps at the detector itself).</div><div>Which is not necessarily a problem, as long as the tilt axis and tilt angles are specified accordingly.</div><div><br></div><div>Best wishes,<br></div><div><div><div dir="ltr" class="gmail_signature" data-smartmail="gmail_signature"><br>--<br>Ricardo Diogo Righetto<br></div></div><br></div></div><br><div class="gmail_quote"><div dir="ltr" class="gmail_attr">Em seg., 18 de mar. de 2024 às 02:57, Sylvain Trepout <<a href="mailto:sylvain.trepout@monash.edu">sylvain.trepout@monash.edu</a>> escreveu:<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="ltr">Hi Zongli and Ricardo, and tomo enthusiasts,<div><br></div><div><div>Same here, on a Krios G4 with Falcon 4i and Tomo software version 5.16.0.6389, the orientation of the tilt axis is not reported correctly.</div><div>TiltAxisAngle = -174.72</div><div>RotationAngle = 84.72</div><div><br></div><div><div>But the correct tilt axis angle found by IMOD is 95.07</div><div>It does not fit the formula you gave Ricardo though... off by just 10 degrees.</div><div>Or maybe this is a flip and then the orientation would be ok, but this would change the handedness.<br></div></div><div><br></div><div>For comparison, on the Krios G1 (with a K3 and an older version of Tomo software)</div><div><div>TiltAxisAngle = -174.47</div><div>RotationAngle = -84.72</div></div><div>The correct tilt axis orientation (as found by IMOD) is around 87 (this fits your formula, Ricardo).</div><div><br></div><div>Best wishes,</div><div>Sylvain</div><div><div><div dir="ltr" class="gmail_signature"><div dir="ltr"><p style="color:rgb(34,34,34);margin-bottom:0px"><span style="color:rgb(80,80,80);font-family:sans-serif">PhD</span><br>Research Fellow - Microscopy Australia</p><p style="color:rgb(34,34,34);margin-bottom:0px"><b>Monash University</b><br></p><span style="color:rgb(34,34,34)">Ramaciotti Centre for Cryo-EM</span><br style="color:rgb(34,34,34)"><span style="color:rgb(34,34,34)">G</span><span style="color:rgb(34,34,34)">92, 15 Innovation Walk, Clayton Campus</span><br style="color:rgb(34,34,34)"><span style="color:rgb(34,34,34)">Victoria, 3800 Australia</span><p style="color:rgb(80,80,80);font-family:sans-serif;margin-bottom:0px">E: <a href="mailto:sylvain.trepout@monash.edu" style="color:rgb(17,85,204)" target="_blank">sylvain.trepout@monash.edu</a></p></div></div></div><br></div></div></div><br><div class="gmail_quote"><div dir="ltr" class="gmail_attr">On Sun, 17 Mar 2024 at 07:53, Zongli Li <<a href="mailto:zongli.li@gmail.com" target="_blank">zongli.li@gmail.com</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="ltr">Dear Ricardo,<div><br></div><div>Thank you so much! This makes a lot of sense.</div><div>Best wishes,</div><div><br></div><div>Zongli</div></div><br><div class="gmail_quote"><div dir="ltr" class="gmail_attr">On Sat, Mar 16, 2024 at 12:37 PM Ricardo Righetto <<a href="mailto:ricardorighetto@gmail.com" target="_blank">ricardorighetto@gmail.com</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="ltr"><div>Dear Zongli,</div><div><br></div><div>Thanks for bringing this up. As far as I know this is a bug (?) in Tomo5. This is how you convert the TiltAxisAngle from the Tomo5 mdoc to match the IMOD/SerialEM convention (followed by RELION and other packages):</div><div><br></div><div>
TiltAxisAngle_SerialEM =
TiltAxisAngle_Tomo5 * (-1) - 90 <br></div><div><br></div><div>Thanks to Grigory Sharov for reporting this conversion previously on the EMAN2 mailing list:</div><div><a href="https://urldefense.com/v3/__https://groups.google.com/g/eman2/c/piux0GdX_7M/m/ScQNh9TNAQAJ__;!!Mih3wA!Go-c8e-QwJVBsU0qttyzgSDv-knvzqN6KJbTdN11SKI-RKgQ9dktWrwtqL_J9HC10IqJx5KxDD2MNvPZcrYB$" target="_blank">https://groups.google.com/g/eman2/c/piux0GdX_7M/m/ScQNh9TNAQAJ</a></div><div><br></div><div>TiltAxisAngle and RotationAngle should always be 90 degrees apart in any mdoc:</div><div><a href="https://urldefense.com/v3/__https://bio3d.colorado.edu/SerialEM/hlp/html/about_formats.htm__;!!Mih3wA!Go-c8e-QwJVBsU0qttyzgSDv-knvzqN6KJbTdN11SKI-RKgQ9dktWrwtqL_J9HC10IqJx5KxDD2MNiJtThea$" target="_blank">https://bio3d.colorado.edu/SerialEM/hlp/html/about_formats.htm</a></div><div>That's why you got the "correct" result when using the value from RotationAngle as the tilt axis. Beware though that you may have inverted handedness in your reconstructions. In my experience, the conversion above should give the correct results.</div><div><br></div><div>Would be great if others can comment if this is indeed the case, and why this bug persists for so long...</div><div><br></div><div>Best wishes,</div><div><div><div dir="ltr" class="gmail_signature"><br>--<br>Ricardo Diogo Righetto<br></div></div><br></div></div><br><div class="gmail_quote"><div dir="ltr" class="gmail_attr">Em sex., 15 de mar. de 2024 às 21:33, Zongli Li <<a href="mailto:zongli.li@gmail.com" target="_blank">zongli.li@gmail.com</a>> escreveu:<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="ltr"><div>Dear colleagues, <br></div><div><br></div><div>We recently collected some tilt series data on vitrified ribosome sample using ThermoFisher Scientifc's Tomo5 software. The data were collected on Falcon 4i. When trying to process the data using the new version of relion tomo workflow (relion 5.0-beta), we are wondering what parameter we should use from the dmoc file for tilt axis angle (TiltAxisAngle or RotationAngle). If the TiltAxisAngle is used in our test run, the reconstructed tomogram is nonsense, but the reconstructed tomogram looks okay if RotationAngle is used. Any suggestions or explanations on this would be highly appreciated.</div><div><br></div><div>Best,</div><div><br></div><div>Zongli<br></div></div>
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