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<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri",sans-serif;color:black">Hi Maarten, I hope the science treats you well.<o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri",sans-serif;color:black"> <o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri",sans-serif;color:black">When I get a registration result that works I feel the overlap in the BDV looks good, although this is a little subjective. Can I ask what kind or errors you get
in the log during the registration? I’ve found that I can get it down to less than a pixel and it looks ok.*, **<o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri",sans-serif;color:black"><o:p> </o:p></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri",sans-serif;color:black">I find that the allowed pixel error helps (I use 1 px), and the significance is less sensitive, but I’ve found 10-15 appears better. Following that the lambda
value is better when higher as far as I can tell. I started to look into the literature, but these things are not so accessible on just skimming the papers.<o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri",sans-serif;color:black"><o:p> </o:p></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri",sans-serif;color:black">I don’t know how much the transformation model and model to regularize with affects things as I haven’t played with those much. Also, I don’t know how the sigma
values and thresholds of the bead detection affect the registration, if at all.<o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri",sans-serif;color:black"><o:p> </o:p></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri",sans-serif;color:black">Thanks<o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri",sans-serif;color:black">Neil<o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri",sans-serif;color:black"> <o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri",sans-serif;color:black">* I have a camera that is not properly sampling. I’ve opted to get a larger area and sacrifice resolution on the old ORCA’s we’re using. I have a pixel spacing
of 0.64 um, which is approximately the resolution with the 20XW lens. Can I ask if anybody knows how my resolution is affected? I assumed that doubling the field of view in this way would leave me with a resolution ~ 1um…?<o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri",sans-serif;color:black">** My calibration samples, registrations, and
<span class="spelle">MVdecons</span> are looking pretty nice, but I’m struggling with the real world samples for some reason… I topic for another post once I’ve tried a few things.<o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-size:11.0pt;font-family:"Calibri",sans-serif;color:black"> <o:p></o:p></span></p>
<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-size:11.0pt;font-family:"Calibri",sans-serif;color:black"> <o:p></o:p></span></p>
<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-size:11.0pt;font-family:"Calibri",sans-serif;color:black"> </span><span style="font-family:"Calibri",sans-serif;color:black"><o:p></o:p></span></p>
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<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><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"> OpenSPIM
[mailto:openspim-bounces@openspim.org] <b>On Behalf Of </b>Maarten Hilbrant<br>
<b>Sent:</b> Monday, August 22, 2016 9:26 AM<br>
<b>To:</b> openspim@openspim.org<br>
<b>Subject:</b> [OpenSPIM] Multiview Reconstruction- optimizing settings?</span><span style="font-family:"Calibri",sans-serif;color:black"><o:p></o:p></span></p>
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<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-family:"Calibri",sans-serif;color:black"> <o:p></o:p></span></p>
<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-family:"Calibri",sans-serif;color:black">Dear all,<o:p></o:p></span></p>
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<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-family:"Calibri",sans-serif;color:black"> <o:p></o:p></span></p>
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<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-family:"Calibri",sans-serif;color:black">nice to see some questions on the mailing list lately regarding Stephan Preibisch' multview plugin. After attending the
EMBO workshop in Dresden in 2014, I've been using the plugin for two years now and it is an essential part of our light sheet workflow. In a nutshell, we image insect embryos, embedded in agarose containing sub-resolution beads, using single-sided illumination
with 8 or 16 angles per time point. In order to infer the biological relevance of RNAi phenotypes, we tend to look at many different specimens for few time points, rather than collecting time-lapse data for a small number of specimens. In practice, this means
a lot of reconstructing and adjusting settings, although I find that, for most of our datasets, default settings work nicely for bead detection, registration and deconvolution ("Nicely" in this case meaning a significant increase in local contrast). Sometimes,
however, registration is sub-optimal (and who knows, maybe it is also for the analyses that I normally quality as "nice''-). In these cases, as a first hint, inspection with the BigDataViewer reveals that the psf's of the different angles don't overlap properly.<o:p></o:p></span></p>
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<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-family:"Calibri",sans-serif;color:black"> <o:p></o:p></span></p>
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<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-family:"Calibri",sans-serif;color:black">- Does this sound familiar to other users of the plugin?<o:p></o:p></span></p>
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<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-family:"Calibri",sans-serif;color:black">- What reconstruction parameters do you generally tinker with first?<o:p></o:p></span></p>
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<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-family:"Calibri",sans-serif;color:black">- for example: do you spend a lot of time on bead detection (and subsequent exclusion)?<o:p></o:p></span></p>
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<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-family:"Calibri",sans-serif;color:black">- Do you sometimes reject datasets for multi view reconstruction and if so, on what grounds? For example, I like to image
"a lot" of agarose surrounding the embryo in order to have a large set of beads for reconstruction, but am not always very systematic about this. Is this crucial in your data acquisition?<o:p></o:p></span></p>
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<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-family:"Calibri",sans-serif;color:black"> <o:p></o:p></span></p>
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<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-family:"Calibri",sans-serif;color:black">any ideas appreciated, basically to make better use of this great tool.<o:p></o:p></span></p>
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<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-family:"Calibri",sans-serif;color:black"> <o:p></o:p></span></p>
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<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-family:"Calibri",sans-serif;color:black">cheers,<o:p></o:p></span></p>
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<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-family:"Calibri",sans-serif;color:black">Maarten<o:p></o:p></span></p>
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<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-family:"Calibri",sans-serif;color:black"> <o:p></o:p></span></p>
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<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-size:13.5pt;font-family:"Helvetica",sans-serif;color:black">******************************<br>
Dr. Maarten Hilbrant<br>
Postdoctoral Researcher<br>
Zoological Institute<br>
University of Cologne<br>
Cologne Biocenter<br>
Zülpicher Str. 47b<br>
50674 Cologne<br>
Germany</span><span style="font-family:"Calibri",sans-serif;color:black"><o:p></o:p></span></p>
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<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-size:13.5pt;font-family:"Helvetica",sans-serif;color:black">+49(0)221-4703238</span><span style="font-family:"Calibri",sans-serif;color:black"><o:p></o:p></span></p>
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<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-family:"Calibri",sans-serif;color:black"><a href="mailto:m.hilbrant@uni-koeln.de"><span style="font-size:13.5pt;font-family:"Helvetica",sans-serif">m.hilbrant@uni-koeln.de</span></a><o:p></o:p></span></p>
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<p class="MsoNormal"><span style="font-family:"Calibri",sans-serif;color:black"><o:p> </o:p></span></p>
<p class="MsoNormal" style="mso-margin-top-alt:auto;mso-margin-bottom-alt:auto"><span style="font-family:"Calibri",sans-serif;color:black"> <o:p></o:p></span></p>
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