<html><head><meta http-equiv="Content-Type" content="text/html charset=windows-1252"></head><body style="word-wrap: break-word; -webkit-nbsp-mode: space; -webkit-line-break: after-white-space;" class="">No worries,<div class=""><br class=""></div><div class="">"<font face="Calibri, sans-serif" class=""><span style="font-size: 14px;" class="">As I understand it SPIM improves the axial resolution of images but does not change lateral resolution relative to widefield. Is this correct?” Correct. It is in essence a widefield microscope system with optical sectioning in the axial direction. </span></font><br class=""><div apple-content-edited="true" class="">
<div style="color: rgb(0, 0, 0); font-family: Helvetica; font-size: 12px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: 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=""><div class=""><br class="Apple-interchange-newline"><br class=""></div><div class="">Best Regards,</div><div class="">Pete</div><div class=""><br class=""></div><div class="">Peter Gabriel Pitrone - DipRMS TechRMS FRMS</div><div class="">Light Sheet Fluorescence Microscopist and Imaging Specialist</div><div class="">for Dr. Pavel Tomancak's research group at the</div><div class="">Max Planck Institute of Molecular Cell Biology and Genetics</div><div class="">Pfotenhauerstraße 108</div><div class="">01307 Dresden, Saxony</div><div class="">Germany</div><div class=""><br class=""></div><div class=""><br class=""></div><div class=""><a href="http://www.mpi-cbg.de/research/research-groups/pavel-tomancak.html" class="">http://www.mpi-cbg.de/research/research-groups/pavel-tomancak.html</a> &</div><div class=""><a href="http://www.openspim.org" class="">http://www.openspim.org</a></div><div class=""><br class=""></div><div class="">"I KEEP six honest serving-men (they taught me all I knew); their names are What and Why and When and How and Where and Who." Rudyard Kipling - The Elephant's Child</div></div><div style="color: rgb(0, 0, 0); font-family: Helvetica; font-size: 12px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: 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=""><br class=""></div><br class="Apple-interchange-newline">
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<br class=""><div><blockquote type="cite" class=""><div class="">On Sep 23, 2015, at 5:20 PM, Feinstein, Timothy N <<a href="mailto:tnf8@pitt.edu" class="">tnf8@pitt.edu</a>> wrote:</div><br class="Apple-interchange-newline"><div class=""><div style="font-family: Calibri, sans-serif; font-size: 14px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: 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=""><div class=""><div class="">Thank you Fredrik and Peter! Right now I am looking at either an Olympus U-AEC 1x/2x magnification changer or else the DE20BXC 2x magnifying camera adapter from SPOT Imaging. </div><div class=""><br class=""></div><div class="">I checked the Nyquist sampling frequency for deconvolution at<span class="Apple-converted-space"> </span><a href="http://www.svi.nl/NyquistCalculator" style="color: purple; text-decoration: underline;" class="">www.svi.nl/NyquistCalculator</a>, and it looks like magnification is a good idea. The optimal sampling frequency for widefield with a 0.5 NA water lens is 0.26 um per px. I am imaging at 0.33 um/px with a 1x camera adapter. Therefore I have a little resolution to gain, though a 2x adapter at 0.16 um/px will be grossly oversampling. It might be a good idea to use a 1.5x adapter instead, as 0.22 um/px is pretty close to the theoretical limit. As I understand it SPIM improves the axial resolution of images but does not change lateral resolution relative to widefield. Is this correct? </div><div class=""><br class=""></div><div class="">All the best, </div><div class=""><br class=""></div><div class=""><br class=""></div><div class="">Tim</div><div class=""><br class=""></div><div class=""><div class="">Timothy Feinstein, Ph.D. </div><div class="">Research Scientist</div><div class="">University of Pittsburgh Department of Developmental Biology</div><div class=""><br class=""></div></div></div></div><div style="font-family: Calibri, sans-serif; font-size: 14px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: 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=""><br class=""></div><span id="OLK_SRC_BODY_SECTION" style="font-family: Calibri, sans-serif; font-size: 14px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: 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=""><div style="font-family: Calibri; font-size: 11pt; text-align: left; border-width: 1pt medium medium; border-style: solid none none; padding: 3pt 0in 0in; border-top-color: rgb(181, 196, 223);" class=""><span style="font-weight: bold;" class="">From:<span class="Apple-converted-space"> </span></span>Fredrik Ek <<a href="mailto:fredrik.ek@med.lu.se" style="color: purple; text-decoration: underline;" class="">fredrik.ek@med.lu.se</a>><br class=""><span style="font-weight: bold;" class="">Date:<span class="Apple-converted-space"> </span></span>Wednesday, September 23, 2015 at 4:34 AM<br class=""><span style="font-weight: bold;" class="">To:<span class="Apple-converted-space"> </span></span>Timothy Feinstein <<a href="mailto:tnf8@pitt.edu" style="color: purple; text-decoration: underline;" class="">tnf8@pitt.edu</a>>, openspim <<a href="mailto:openspim@openspim.org" style="color: purple; text-decoration: underline;" class="">openspim@openspim.org</a>><br class=""><span style="font-weight: bold;" class="">Subject:<span class="Apple-converted-space"> </span></span>SV: changing "objectives"<br class=""></div><div class=""><br class=""></div><div xmlns:v="urn:schemas-microsoft-com:vml" xmlns:o="urn:schemas-microsoft-com:office:office" xmlns:w="urn:schemas-microsoft-com:office:word" xmlns:m="http://schemas.microsoft.com/office/2004/12/omml" xmlns="http://www.w3.org/TR/REC-html40" class=""><div lang="EN-US" link="blue" vlink="purple" class=""><div class="WordSection1" style="page: WordSection1;"><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 11pt; font-family: Calibri, sans-serif; color: rgb(31, 73, 125);" class="">Hi,<o:p class=""></o:p></span></div><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 11pt; font-family: Calibri, sans-serif; color: rgb(31, 73, 125);" class=""><o:p class=""> </o:p></span></div><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 11pt; font-family: Calibri, sans-serif; color: rgb(31, 73, 125);" class="">We are using a Olympus magnifier 1x,2x and 4x with c-mount together with a Flash4.0 v2 and it works very well with the SPIM setup (20x 0.5 objective).<o:p class=""></o:p></span></div><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 11pt; font-family: Calibri, sans-serif; color: rgb(31, 73, 125);" class=""><o:p class=""> </o:p></span></div><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 11pt; font-family: Calibri, sans-serif; color: rgb(31, 73, 125);" class="">Best,<o:p class=""></o:p></span></div><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 11pt; font-family: Calibri, sans-serif; color: rgb(31, 73, 125);" class=""><o:p class=""> </o:p></span></div><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 11pt; font-family: Calibri, sans-serif; color: rgb(31, 73, 125);" class="">Fredrik<o:p class=""></o:p></span></div><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10pt; font-family: Arial, sans-serif;" class=""><o:p class=""> </o:p></span></div><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10pt; font-family: Arial, sans-serif;" class="">Fredrik Ek, PhD,</span><span style="font-size: 10pt; font-family: Tahoma, sans-serif;" class=""><o:p class=""></o:p></span></div><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10pt; font-family: Arial, sans-serif;" class="">Researcher, Chemical Biology & Therapeutics<span class="Apple-converted-space"> </span></span><span style="font-size: 10pt; font-family: Tahoma, sans-serif;" class=""><br class=""></span><span style="font-size: 10pt; font-family: Arial, sans-serif;" class="">Department of Experimental Medical Science<span class="Apple-converted-space"> </span></span><span style="font-size: 10pt; font-family: Tahoma, sans-serif;" class=""><br class=""></span><span style="font-size: 10pt; font-family: Arial, sans-serif;" class="">Lund University, BMC D10<span class="Apple-converted-space"> </span></span><span style="font-size: 10pt; font-family: Tahoma, sans-serif;" class=""><br class=""></span><span style="font-size: 10pt; font-family: Arial, sans-serif;" class="">S-221 84 Lund, SWEDEN<span class="Apple-converted-space"> </span></span><span style="font-size: 10pt; font-family: Tahoma, sans-serif;" class=""><br class=""></span><span style="font-size: 10pt; font-family: Arial, sans-serif;" class="">------------------------<span class="Apple-converted-space"> </span></span><span style="font-size: 10pt; font-family: Tahoma, sans-serif;" class=""><br class=""></span><span lang="SV" style="font-size: 10pt; font-family: Tahoma, sans-serif;" class=""><a href="mailto:Fredrik.Ek@med.lu.se" style="color: purple; text-decoration: underline;" class=""><span lang="EN-US" style="font-family: Arial, sans-serif; color: blue;" class="">Fredrik.Ek@med.lu.se</span></a></span><span lang="SV" style="font-size: 10pt; font-family: Arial, sans-serif;" class=""></span><span style="font-size: 10pt; font-family: Tahoma, sans-serif;" class=""><br class=""></span><span style="font-size: 10pt; font-family: Arial, sans-serif;" class="">Phone: +46-727 17 22 05</span><span style="font-size: 10pt; font-family: Tahoma, sans-serif;" class=""><o:p class=""></o:p></span></div><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 11pt; font-family: Calibri, sans-serif; color: rgb(31, 73, 125);" class=""><o:p class=""> </o:p></span></div><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 11pt; font-family: Calibri, sans-serif; color: rgb(31, 73, 125);" class=""><o:p class=""> </o:p></span></div><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 11pt; font-family: Calibri, sans-serif; color: rgb(31, 73, 125);" class=""><o:p class=""> </o:p></span></div><div class=""><div style="border-style: solid none none; border-top-color: rgb(181, 196, 223); border-top-width: 1pt; padding: 3pt 0cm 0cm;" class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><b class=""><span lang="SV" style="font-size: 10pt; font-family: Tahoma, sans-serif;" class="">Från:</span></b><span lang="SV" style="font-size: 10pt; font-family: Tahoma, sans-serif;" class=""><span class="Apple-converted-space"> </span><a href="mailto:openspim-bounces@openspim.org" style="color: purple; text-decoration: underline;" class="">openspim-bounces@openspim.org</a><span class="Apple-converted-space"> </span>[<a href="mailto:openspim-bounces@openspim.org" style="color: purple; text-decoration: underline;" class="">mailto:openspim-bounces@openspim.org</a>]<span class="Apple-converted-space"> </span><b class="">För<span class="Apple-converted-space"> </span></b>Feinstein, Timothy N<br class=""><b class="">Skickat:</b><span class="Apple-converted-space"> </span>den 22 september 2015 17:35<br class=""><b class="">Till:</b><span class="Apple-converted-space"> </span>openspim<br class=""><b class="">Ämne:</b><span class="Apple-converted-space"> </span>[OpenSPIM] changing "objectives"<o:p class=""></o:p></span></div></div></div><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><o:p class=""> </o:p></div><div class=""><div class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10.5pt; font-family: Calibri, sans-serif;" class="">Hi folks, <o:p class=""></o:p></span></div></div><div class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10.5pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></span></div></div><div class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10.5pt; font-family: Calibri, sans-serif;" class="">Thanks to the indispensable help of many users here, our SPIM is up and working. You are all the best. <o:p class=""></o:p></span></div></div><div class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10.5pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></span></div></div><div class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10.5pt; font-family: Calibri, sans-serif;" class="">Since a SPIM makes it quite hard to change objectives, I bought 1x and a 0.5x C-mount camera adapters to get the equivalent of a 20x and a 10x objective. It seems to work quite well. Now I wonder whether anyone has tried a magnifying tube lens like the Olympus U-CA or U-ECA to get a higher power image. It all depends on whether a 'vanilla' openSPIM design is already imaging near the Nyquist limit. We get 0.33 um/px using the Olympus 20x/0.5 and a Hamamatsu Flash4.0 v2, and even with deconvolution it may be pointless to magnify the image any further. Any thoughts appreciated. <o:p class=""></o:p></span></div></div><div class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10.5pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></span></div></div><div class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10.5pt; font-family: Calibri, sans-serif;" class="">Alternatively, has anyone tried keeping a second sample chamber with a higher power objective to swap in as needed? It seems like a pain but less cumbersome than building a second instrument. <o:p class=""></o:p></span></div></div><div class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10.5pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></span></div></div><div class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10.5pt; font-family: Calibri, sans-serif;" class="">Thanks, <o:p class=""></o:p></span></div></div><div class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10.5pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></span></div></div><div class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10.5pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></span></div></div><div class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10.5pt; font-family: Calibri, sans-serif;" class="">Tim<o:p class=""></o:p></span></div></div><div class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10.5pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></span></div></div><div class=""><div class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10.5pt; font-family: Calibri, sans-serif;" class="">Timothy Feinstein, Ph.D. <o:p class=""></o:p></span></div></div><div class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10.5pt; font-family: Calibri, sans-serif;" class="">Research Scientist<o:p class=""></o:p></span></div></div><div class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10.5pt; font-family: Calibri, sans-serif;" class="">University of Pittsburgh Department of Developmental Biology<o:p class=""></o:p></span></div></div><div class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10.5pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></span></div></div></div></div><div class=""><div style="margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: 'Times New Roman', serif;" class=""><span style="font-size: 10.5pt; font-family: Calibri, sans-serif;" class=""><o:p class=""> </o:p></span></div></div></div></div></div></span><span style="font-family: Calibri, sans-serif; font-size: 14px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: 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><span style="font-family: Calibri, sans-serif; font-size: 14px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: 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: Calibri, sans-serif; font-size: 14px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: 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=""><span style="font-family: Calibri, sans-serif; font-size: 14px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: 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="">OpenSPIM mailing list</span><br style="font-family: Calibri, sans-serif; font-size: 14px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: 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=""><span style="font-family: Calibri, sans-serif; font-size: 14px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: 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=""><a href="mailto:OpenSPIM@openspim.org" class="">OpenSPIM@openspim.org</a></span><br style="font-family: Calibri, sans-serif; font-size: 14px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: 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=""><span style="font-family: Calibri, sans-serif; font-size: 14px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: 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=""><a href="http://openspim.org/mailman/listinfo/openspim" class="">http://openspim.org/mailman/listinfo/openspim</a></span></div></blockquote></div><br class=""></div></body></html>