Zeiss 25 X Plan Neofluor Phase 3 objective for sale
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Goodhouse, Joseph G. |
Zeiss 25 X Plan Neofluor Phase 3 objective for sale
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Dear Microscopists
Please respond directly to me. Regards, Joe Goodhouse
Visit us at http://www.molbio1.princeton.edu/facility/confocal/ |
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Julian Smith III |
Re: Zeiss 25 X Plan Neofluor Phase 3 objective for sale
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Joe
We still have an inverted zeiss fluorescence 'scope. What sort of price is the user looking for? If it's in the $100-$150 range, we're interested. Julian Goodhouse, Joseph G. wrote: > > Dear Microscopists > Is anyone in need of a Zeiss 25 X Plan Neofluor / 0.9 > NA Multiple Immersion Phase 3 objective. [Part # is 461726] This is a > 160 mm tube lens and is in excellent. Owner no longer needs it and is > willing to sell. Price negotiable. > > Please respond directly to me. > > Regards, > > > Joe Goodhouse > Confocal Core Lab Manager > Dept. of Molecular Biology > Princeton University > 609-258-5432 > > Visit us at _http://www.molbio1.princeton.edu/facility/confocal/_ > <http://www.molbio1.princeton.edu/facility/confocal/> > > -- Julian P.S. Smith III Director, Winthrop Microscopy Facility Dept. of Biology Winthrop University 520 Cherry Rd. Rock Hill, SC 29733 803-323-2111 x6427 (vox) 803-323-3448 (fax) 803-524-2347 (cell) |
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When acquiring confocal data, we find it useful to have a previous experiment
(usually an image acquired earlier during that session) open, while making the live acquisition. This allows an immediate visual comparison of the current result with a previous result. On the Leica SP5, we do not seem to be able to do this, because apparently only one data window can be opened, and if a new scan is done, it replaces the old data in that window. I suppose I can use ImageJ to open the stored data, but i was wondering if we are missing something obvious. --aryeh -- Aryeh Weiss School of Engineering Bar Ilan University Ramat Gan 52900 Israel Ph: 972-3-5317638 FAX: 972-3-7384050 |
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Aryeh,
I hope this helps, Zoltan On Mon, Feb 9, 2009 at 10:31 PM, Aryeh Weiss <[hidden email]> wrote: When acquiring confocal data, we find it useful to have a previous experiment (usually an image acquired earlier during that session) open, while making the live acquisition. This allows an immediate visual comparison of the current result with a previous result. On the Leica SP5, we do not seem to be able to do this, because apparently only one data window can be opened, and if a new scan is done, it replaces the old data in that window. -- Zoltan Cseresnyes Facility manager, Imaging Suite University of Cambridge, UK
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Hello Zoltan & listers,
You comments suggest that you are a regular user of
the Leica Live Data Mode on the SP5... if so, I was wondering how you handle the
data post-acquisition - to get back to individual time series from each of
the different positions. We tend to use ImageJ and macros we have written
to sort and concatenate the files, but I am interested in how others approach
this.
Best wishes,
Graham
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Also, does the Mark & Find function work in Live Data in version 2? When I had a quick look at in 1.8.2 it didn't seem like "mark and find" was available in Live Data mode? (or at least I couldn't see it). Regards, Adrian Smith Centenary Institute, Sydney, Australia On 10/02/2009, at 2:32 PM, Dr. Graham Wright wrote:
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First -- thanks for the quick responses. We will ask our rep to get us the
software upgrade (the system is only a few months old -- I assume that getting the software will not be a problem). Concerning mark and find with live data mode, I think that it is not available on our machine. This is a problem because we have not been able to find a way to do a multipoint timelapse acquisition. Maybe the experienced SP5 users can correct me on that one. Is there a way to set up an XYZT acquisition with multiple fields? --aryeh Adrian Smith wrote: > Also, does the Mark & Find function work in Live Data in version 2? > > When I had a quick look at in 1.8.2 it didn't seem like "mark and find" > was available in Live Data mode? (or at least I couldn't see it). > > Regards, > > Adrian Smith > Centenary Institute, Sydney, Australia > > > > On 10/02/2009, at 2:32 PM, Dr. Graham Wright wrote: > >> Hello Zoltan & listers, >> >> You comments suggest that you are a regular user of the Leica Live >> Data Mode on the SP5... if so, I was wondering how you handle the data >> post-acquisition - to get back to individual time series from each of >> the different positions. We tend to use ImageJ and macros we have >> written to sort and concatenate the files, but I am interested in how >> others approach this. >> >> Best wishes, >> Graham >> >> ----- Original Message ----- >> *From:* Zoltan Cseresnyes <mailto:[hidden email]> >> *To:* [hidden email] >> <mailto:[hidden email]> >> *Sent:* Tuesday, February 10, 2009 6:36 AM >> *Subject:* Re: Leica SP5 question >> >> Aryeh, >> >> If you upgrade to LAS AF version 2.0, you'll be able to open a >> second image window and observe previously acquired data. We very >> much like the new version anyway, e.g. we find the Mark and Find >> and Live Data Mode more stable than even with the 1.8.2, which was >> already pretty good. >> I hope this helps, >> >> Zoltan >> >> >> >> On Mon, Feb 9, 2009 at 10:31 PM, Aryeh Weiss <[hidden email] >> <mailto:[hidden email]>> wrote: >> >> When acquiring confocal data, we find it useful to have a >> previous experiment (usually an image acquired earlier during >> that session) open, while making the live acquisition. This >> allows an immediate visual comparison of the current result >> with a previous result. On the Leica SP5, we do not seem to be >> able to do this, because apparently only one data window can >> be opened, and if a new scan is done, it replaces the old data >> in that window. >> >> I suppose I can use ImageJ to open the stored data, but i was >> wondering if we are missing something obvious. >> >> --aryeh >> -- >> Aryeh Weiss >> School of Engineering >> Bar Ilan University >> Ramat Gan 52900 Israel >> >> Ph: 972-3-5317638 >> FAX: 972-3-7384050 >> >> >> >> >> -- >> >> Zoltan Cseresnyes >> Facility manager, Imaging Suite >> University of Cambridge, UK > |
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Hi Andy,
To be able to do XYZT acquisition with multiple fields your scope needs to be fitted with a motorised stage. If you have that then you will be fine. To set up mark your first position, give it a z range and press the exchange position button. If you don't hit the exchange position button all your z stacks will be based on the last settings you give it. I have done XYZT experiments in the past that have been 10-15 Z planes over 24 different stage positions. Cheers Cam Cameron J. Nowell Microscopy Manager Centre for Advanced Microscopy Ludwig Institute for Cancer Research PO Box 2008 Royal Melbourne Hospital Victoria, 3050 AUSTRALIA Office: +61 3 9341 3155 Mobile: +61422882700 Fax: +61 3 9341 3104 Facility Website -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Aryeh Weiss Sent: Tuesday, 10 February 2009 4:47 PM To: [hidden email] Subject: Re: Leica SP5 question First -- thanks for the quick responses. We will ask our rep to get us the software upgrade (the system is only a few months old -- I assume that getting the software will not be a problem). Concerning mark and find with live data mode, I think that it is not available on our machine. This is a problem because we have not been able to find a way to do a multipoint timelapse acquisition. Maybe the experienced SP5 users can correct me on that one. Is there a way to set up an XYZT acquisition with multiple fields? --aryeh Adrian Smith wrote: > Also, does the Mark & Find function work in Live Data in version 2? > > When I had a quick look at in 1.8.2 it didn't seem like "mark and find" > was available in Live Data mode? (or at least I couldn't see it). > > Regards, > > Adrian Smith > Centenary Institute, Sydney, Australia > > > > On 10/02/2009, at 2:32 PM, Dr. Graham Wright wrote: > >> Hello Zoltan & listers, >> >> You comments suggest that you are a regular user of the Leica Live >> Data Mode on the SP5... if so, I was wondering how you handle the >> post-acquisition - to get back to individual time series from each of >> the different positions. We tend to use ImageJ and macros we have >> written to sort and concatenate the files, but I am interested in how >> others approach this. >> >> Best wishes, >> Graham >> >> ----- Original Message ----- >> *From:* Zoltan Cseresnyes <mailto:[hidden email]> >> *To:* [hidden email] >> <mailto:[hidden email]> >> *Sent:* Tuesday, February 10, 2009 6:36 AM >> *Subject:* Re: Leica SP5 question >> >> Aryeh, >> >> If you upgrade to LAS AF version 2.0, you'll be able to open a >> second image window and observe previously acquired data. We very >> much like the new version anyway, e.g. we find the Mark and Find >> and Live Data Mode more stable than even with the 1.8.2, which >> already pretty good. >> I hope this helps, >> >> Zoltan >> >> >> >> On Mon, Feb 9, 2009 at 10:31 PM, Aryeh Weiss <[hidden email] >> <mailto:[hidden email]>> wrote: >> >> When acquiring confocal data, we find it useful to have a >> previous experiment (usually an image acquired earlier during >> that session) open, while making the live acquisition. This >> allows an immediate visual comparison of the current result >> with a previous result. On the Leica SP5, we do not seem to >> able to do this, because apparently only one data window can >> be opened, and if a new scan is done, it replaces the old data >> in that window. >> >> I suppose I can use ImageJ to open the stored data, but i was >> wondering if we are missing something obvious. >> >> --aryeh >> -- >> Aryeh Weiss >> School of Engineering >> Bar Ilan University >> Ramat Gan 52900 Israel >> >> Ph: 972-3-5317638 >> FAX: 972-3-7384050 >> >> >> >> >> -- >> >> Zoltan Cseresnyes >> Facility manager, Imaging Suite >> University of Cambridge, UK > No virus found in this incoming message. Checked by AVG - www.avg.com Version: 8.0.233 / Virus Database: 270.10.19/1942 - Release Date: 02/09/09 17:40:00 This communication is intended only for the named recipient and may contain information that is confidential, legally privileged or subject to copyright; the Ludwig Institute for Cancer Research Ltd does not waiver any rights if you have received this communication in error. The views expressed in this communication are those of the sender and do not necessarily reflect the views of the Ludwig Institute for Cancer Research Ltd. |
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In reply to this post by Aryeh Weiss
Dear Aryeh
I have some experience with Leica Sp5 and despite my attempts I was not able to find a way of having more than one image open at the same time so that comparisons of images in acquisition with previous images or even side by side comparisons of two images was very arduous. Any one knows if there are plans from Leica to fix this? Cordially Caterina ____________________________ Caterina Strambio De Castillia, Dottore in Biologia, PhD Senior Research Scientist Laboratory of Viral replication, pathogenesis, and immunity http://www.mimo.unige.ch/luban_lab/ Department of Microbiology and Molecular Medicine University of Geneva Switzerland Phone: +41 (0)78 7367537 **** Visiting Scientist Laboratory of Cellular and Structural Biology The Rockefeller University 1230 York Avenue New York NY 10065 On 9-feb-09, at 23:31, Aryeh Weiss wrote: > When acquiring confocal data, we find it useful to have a previous > experiment (usually an image acquired earlier during that session) > open, while making the live acquisition. This allows an immediate > visual comparison of the current result with a previous result. On > the Leica SP5, we do not seem to be able to do this, because > apparently only one data window can be opened, and if a new scan is > done, it replaces the old data in that window. > > I suppose I can use ImageJ to open the stored data, but i was > wondering if we are missing something obvious. > > --aryeh > -- > Aryeh Weiss > School of Engineering > Bar Ilan University > Ramat Gan 52900 Israel > > Ph: 972-3-5317638 > FAX: 972-3-7384050 |
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In reply to this post by Zoltan
There you go!
Caterina ____________________________ Caterina Strambio De Castillia, Dottore in Biologia, PhD Senior Research Scientist Laboratory of Viral replication, pathogenesis, and immunity http://www.mimo.unige.ch/luban_lab/ Department of Microbiology and Molecular Medicine University of Geneva Switzerland Phone: +41 (0)78 7367537 **** Visiting Scientist Laboratory of Cellular and Structural Biology The Rockefeller University 1230 York Avenue New York NY 10065 On 9-feb-09, at 23:36, Zoltan Cseresnyes wrote: > Aryeh, > > If you upgrade to LAS AF version 2.0, you'll be able to open a > second image window and observe previously acquired data. We very > much like the new version anyway, e.g. we find the Mark and Find > and Live Data Mode more stable than even with the 1.8.2, which was > already pretty good. > I hope this helps, > > Zoltan > > > > On Mon, Feb 9, 2009 at 10:31 PM, Aryeh Weiss <[hidden email]> > wrote: > When acquiring confocal data, we find it useful to have a previous > experiment (usually an image acquired earlier during that session) > open, while making the live acquisition. This allows an immediate > visual comparison of the current result with a previous result. On > the Leica SP5, we do not seem to be able to do this, because > apparently only one data window can be opened, and if a new scan is > done, it replaces the old data in that window. > > I suppose I can use ImageJ to open the stored data, but i was > wondering if we are missing something obvious. > > --aryeh > -- > Aryeh Weiss > School of Engineering > Bar Ilan University > Ramat Gan 52900 Israel > > Ph: 972-3-5317638 > FAX: 972-3-7384050 > > > > -- > > Zoltan Cseresnyes > Facility manager, Imaging Suite > University of Cambridge, UK |
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SUBSCRIBE CONFOCALMICROSCOPY Anonymous |
AW: Leica SP5 question
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In reply to this post by Caterina Strambio
Thorsten Staudt
German Cancer Research Center (DKFZ) Department of High Resolution Optical Microscopy (Prof. S. W. Hell) INF 280 69120 Heidelberg, Germany PHONE: +49-6221-51421 -----Ursprüngliche Nachricht----- Von: Confocal Microscopy List im Auftrag von Caterina Strambio De Castillia Gesendet: Di 10.02.2009 11:21 An: [hidden email] Betreff: Re: Leica SP5 question Dear Aryeh I have some experience with Leica Sp5 and despite my attempts I was not able to find a way of having more than one image open at the same time so that comparisons of images in acquisition with previous images or even side by side comparisons of two images was very arduous. Any one knows if there are plans from Leica to fix this? Cordially Caterina ____________________________ Caterina Strambio De Castillia, Dottore in Biologia, PhD Senior Research Scientist Laboratory of Viral replication, pathogenesis, and immunity http://www.mimo.unige.ch/luban_lab/ Department of Microbiology and Molecular Medicine University of Geneva Switzerland Phone: +41 (0)78 7367537 **** Visiting Scientist Laboratory of Cellular and Structural Biology The Rockefeller University 1230 York Avenue New York NY 10065 On 9-feb-09, at 23:31, Aryeh Weiss wrote: > When acquiring confocal data, we find it useful to have a previous > experiment (usually an image acquired earlier during that session) > open, while making the live acquisition. This allows an immediate > visual comparison of the current result with a previous result. On > the Leica SP5, we do not seem to be able to do this, because > apparently only one data window can be opened, and if a new scan is > done, it replaces the old data in that window. > > I suppose I can use ImageJ to open the stored data, but i was > wondering if we are missing something obvious. > > --aryeh > -- > Aryeh Weiss > School of Engineering > Bar Ilan University > Ramat Gan 52900 Israel > > Ph: 972-3-5317638 > FAX: 972-3-7384050 |
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In reply to this post by Cameron Nowell
Hi, Aryeh,
As Cam said below, to do an XYZT on multiple fields you need a motorized stage. Also, when the LAS AF boots up, you have to select "Yes" when the application asks you if you want to initialize the Motorized Stage. Even though the stage initializes when the microscope is turned on, you have to initialize it again in order to load the tile scan and the multipoint capabilities. Hope this helps, George George Ring, Ph.D. Dept. of Cell and Developmental Biology SUNY Upstate Medical University 750 E. Adams St. Syracuse NY 13210 Tel. (315) 464-8595 FAX (315) 464-8535 email: [hidden email] >>> On Tue, Feb 10, 2009 at 12:59 AM, in message <[hidden email]>, Cameron Nowell <[hidden email]> wrote: > Hi Andy, > To be able to do XYZT acquisition with multiple fields your > scope needs to be fitted with a motorised stage. If you have that then > you will be fine. > > To set up mark your first position, give it a z range and press the > exchange position button. If you don't hit the exchange position button > all your z stacks will be based on the last settings you give it. > > I have done XYZT experiments in the past that have been 10-15 Z planes > over 24 different stage positions. > > > Cheers > > > Cam > > > > > Cameron J. Nowell > Microscopy Manager > Centre for Advanced Microscopy > Ludwig Institute for Cancer Research > PO Box 2008 > Royal Melbourne Hospital > Victoria, 3050 > AUSTRALIA > Office: +61 3 9341 3155 > Mobile: +61422882700 > Fax: +61 3 9341 3104 > Facility Website > > > > -----Original Message----- > From: Confocal Microscopy List [mailto:[hidden email]] > On Behalf Of Aryeh Weiss > Sent: Tuesday, 10 February 2009 4:47 PM > To: [hidden email] > Subject: Re: Leica SP5 question > > First -- thanks for the quick responses. We will ask our rep to get us > the > software upgrade (the system is only a few months old -- I assume that > getting > the software will not be a problem). > > Concerning mark and find with live data mode, > I think that it is not available on our machine. This is a problem > because we > have not been able to find a way to do a multipoint timelapse > acquisition. > > Maybe the experienced SP5 users can correct me on that one. Is there a > way to > set up an XYZT acquisition with multiple fields? > > --aryeh > > Adrian Smith wrote: >> Also, does the Mark & Find function work in Live Data in version 2? >> >> When I had a quick look at in 1.8.2 it didn't seem like "mark and > find" >> was available in Live Data mode? (or at least I couldn't see it). >> >> Regards, >> >> Adrian Smith >> Centenary Institute, Sydney, Australia >> >> >> >> On 10/02/2009, at 2:32 PM, Dr. Graham Wright wrote: >> >>> Hello Zoltan & listers, >>> >>> You comments suggest that you are a regular user of the Leica Live >>> Data Mode on the SP5... if so, I was wondering how you handle the > data >>> post-acquisition - to get back to individual time series from each of > >>> the different positions. We tend to use ImageJ and macros we have >>> written to sort and concatenate the files, but I am interested in how > >>> others approach this. >>> >>> Best wishes, >>> Graham >>> >>> ----- Original Message ----- >>> *From:* Zoltan Cseresnyes <mailto:[hidden email]> >>> *To:* [hidden email] >>> <mailto:[hidden email]> >>> *Sent:* Tuesday, February 10, 2009 6:36 AM >>> *Subject:* Re: Leica SP5 question >>> >>> Aryeh, >>> >>> If you upgrade to LAS AF version 2.0, you'll be able to open a >>> second image window and observe previously acquired data. We very >>> much like the new version anyway, e.g. we find the Mark and Find >>> and Live Data Mode more stable than even with the 1.8.2, which > was >>> already pretty good. >>> I hope this helps, >>> >>> Zoltan >>> >>> >>> >>> On Mon, Feb 9, 2009 at 10:31 PM, Aryeh Weiss <[hidden email] >>> <mailto:[hidden email]>> wrote: >>> >>> When acquiring confocal data, we find it useful to have a >>> previous experiment (usually an image acquired earlier during >>> that session) open, while making the live acquisition. This >>> allows an immediate visual comparison of the current result >>> with a previous result. On the Leica SP5, we do not seem to > be >>> able to do this, because apparently only one data window can >>> be opened, and if a new scan is done, it replaces the old > data >>> in that window. >>> >>> I suppose I can use ImageJ to open the stored data, but i was >>> wondering if we are missing something obvious. >>> >>> --aryeh >>> -- >>> Aryeh Weiss >>> School of Engineering >>> Bar Ilan University >>> Ramat Gan 52900 Israel >>> >>> Ph: 972-3-5317638 >>> FAX: 972-3-7384050 >>> >>> >>> >>> >>> -- >>> >>> Zoltan Cseresnyes >>> Facility manager, Imaging Suite >>> University of Cambridge, UK >> > > No virus found in this incoming message. > Checked by AVG - www.avg.com > Version: 8.0.233 / Virus Database: 270.10.19/1942 - Release Date: > 02/09/09 17:40:00 > > > This communication is intended only for the named recipient and may > information that is confidential, legally privileged or subject to copyright; > the Ludwig Institute for Cancer Research Ltd does not waiver any rights if > you have received this communication in error. > The views expressed in this communication are those of the sender and do not > necessarily reflect the views of the Ludwig Institute for Cancer Research > Ltd. |
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In reply to this post by Cameron Nowell
Dear All,
We experience quite visible axial thermal drift with the Prior H117 motorized stage under TIR illumination (ca. 10 nm per second) with the Nikon 100x N.A. 1.45 objective. I have contacted Prior and I was told that they never measured thermal drift for the H117 stage at that level - 100-200 nm. XY motors attached to the bottom of the stage, thus the heat generated by the XY motors could also contribute to the drift. Nikon offers PFS or their Ti50 scope to solve the problem, a kind of expensive solution for the Nikon's first generation, visibly under-engineered TIR system. I would greatly appreciate any positive feedback/suggestions. Thank you, Vitaly NCI-Frederick, 301-846-6575 -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Cameron Nowell Sent: Tuesday, February 10, 2009 1:00 AM To: [hidden email] Subject: Re: Leica SP5 question Hi Andy, To be able to do XYZT acquisition with multiple fields your scope needs to be fitted with a motorised stage. If you have that then you will be fine. To set up mark your first position, give it a z range and press the exchange position button. If you don't hit the exchange position button all your z stacks will be based on the last settings you give it. I have done XYZT experiments in the past that have been 10-15 Z planes over 24 different stage positions. Cheers Cam Cameron J. Nowell Microscopy Manager Centre for Advanced Microscopy Ludwig Institute for Cancer Research PO Box 2008 Royal Melbourne Hospital Victoria, 3050 AUSTRALIA Office: +61 3 9341 3155 Mobile: +61422882700 Fax: +61 3 9341 3104 Facility Website -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Aryeh Weiss Sent: Tuesday, 10 February 2009 4:47 PM To: [hidden email] Subject: Re: Leica SP5 question First -- thanks for the quick responses. We will ask our rep to get us the software upgrade (the system is only a few months old -- I assume that getting the software will not be a problem). Concerning mark and find with live data mode, I think that it is not available on our machine. This is a problem because we have not been able to find a way to do a multipoint timelapse acquisition. Maybe the experienced SP5 users can correct me on that one. Is there a way to set up an XYZT acquisition with multiple fields? --aryeh Adrian Smith wrote: > Also, does the Mark & Find function work in Live Data in version 2? > > When I had a quick look at in 1.8.2 it didn't seem like "mark and find" > was available in Live Data mode? (or at least I couldn't see it). > > Regards, > > Adrian Smith > Centenary Institute, Sydney, Australia > > > > On 10/02/2009, at 2:32 PM, Dr. Graham Wright wrote: > >> Hello Zoltan & listers, >> >> You comments suggest that you are a regular user of the Leica Live >> Data Mode on the SP5... if so, I was wondering how you handle the >> post-acquisition - to get back to individual time series from each of >> the different positions. We tend to use ImageJ and macros we have >> written to sort and concatenate the files, but I am interested in how >> others approach this. >> >> Best wishes, >> Graham >> >> ----- Original Message ----- >> *From:* Zoltan Cseresnyes <mailto:[hidden email]> >> *To:* [hidden email] >> <mailto:[hidden email]> >> *Sent:* Tuesday, February 10, 2009 6:36 AM >> *Subject:* Re: Leica SP5 question >> >> Aryeh, >> >> If you upgrade to LAS AF version 2.0, you'll be able to open a >> second image window and observe previously acquired data. We very >> much like the new version anyway, e.g. we find the Mark and Find >> and Live Data Mode more stable than even with the 1.8.2, which >> already pretty good. >> I hope this helps, >> >> Zoltan >> >> >> >> On Mon, Feb 9, 2009 at 10:31 PM, Aryeh Weiss <[hidden email] >> <mailto:[hidden email]>> wrote: >> >> When acquiring confocal data, we find it useful to have a >> previous experiment (usually an image acquired earlier during >> that session) open, while making the live acquisition. This >> allows an immediate visual comparison of the current result >> with a previous result. On the Leica SP5, we do not seem to >> able to do this, because apparently only one data window can >> be opened, and if a new scan is done, it replaces the old data >> in that window. >> >> I suppose I can use ImageJ to open the stored data, but i was >> wondering if we are missing something obvious. >> >> --aryeh >> -- >> Aryeh Weiss >> School of Engineering >> Bar Ilan University >> Ramat Gan 52900 Israel >> >> Ph: 972-3-5317638 >> FAX: 972-3-7384050 >> >> >> >> >> -- >> >> Zoltan Cseresnyes >> Facility manager, Imaging Suite >> University of Cambridge, UK > No virus found in this incoming message. Checked by AVG - www.avg.com Version: 8.0.233 / Virus Database: 270.10.19/1942 - Release Date: 02/09/09 17:40:00 This communication is intended only for the named recipient and may contain information that is confidential, legally privileged or subject to copyright; the Ludwig Institute for Cancer Research Ltd does not waiver any rights if you have received this communication in error. The views expressed in this communication are those of the sender and do not necessarily reflect the views of the Ludwig Institute for Cancer Research Ltd. |
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Adrian Smith-6 |
Re: Leica SP5 question - Motorised Stage
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In reply to this post by Cameron Nowell
On 10/02/2009, at 4:59 PM, Cameron Nowell wrote:
> To be able to do XYZT acquisition with multiple fields your > scope needs to be fitted with a motorised stage. If you have that then > you will be fine. While we are talking SP5 and motorised stage.... I've been wondering if anyone has a come up with a good procedure for setting up the calibration of the stage position, ie in the Stage Configuration window there are settings for Tile Scan overlap, origin offset sizes, merge settings that need to be set up in order for tile scans to work. One of the students here got them quite close through a laborious trial-and-error process. Unfortunately, we need to reset them because the stage alignment got messed up at one point (when the stage cable got caught in the opening into the incubation enclosure). Before I go through the process of doing it by trial and error or working out a more systematic way of doing it I was wondering if anyone else had any suggestions as to how best approach it? Regards, Adrian Smith Centenary Institute, Sydney, Australia |
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Adrian,
We didn't really have to alter the factory settings in order to get the tiling to work very well (some of my users scan entire locust brains or parts of butterfly wings, which require at least 3x3 tiling, up to 5x5). We had no problem tiling and the LAS AF automatic stitching option worked very well (tiling overlap set to zero). I'm home now but tomorrow I'll try to remember to read the parameters out for you. However, the only critical step was to make sure that the scan field rotation is set such that the sample moves perfectly parallel to the horizontal (X) edge of the screen when we are turning the X controller joystick knob. We select a bright spot in the sample, move it very close to the lower edge of the image screen and slowly turn the joystick knob (X) while watching the bright spot move; we then adjust the field rotation until the bright spot moves perfectly parallel to the lower edge of the screen. In our case this angle turned out to be -0.7 degrees.
I hope this helps, Zoltan On Tue, Feb 10, 2009 at 11:01 PM, Adrian Smith <[hidden email]> wrote: On 10/02/2009, at 4:59 PM, Cameron Nowell wrote: -- Zoltan Cseresnyes Facility manager, Imaging Suite University of Cambridge, UK
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In reply to this post by Graham Wright-5
Hi Graham,
We do it similarly, except that I wrote a C# software toolkit that separates the Leica files into folders based on various criteria, taking advantage of Leica's fairly consistent file-naming structure. Bit of work that extends post-processing time but we learned to live with it.
Best, Zoltan On Tue, Feb 10, 2009 at 3:32 AM, Dr. Graham Wright <[hidden email]> wrote:
-- Zoltan Cseresnyes Facility manager, Imaging Suite University of Cambridge, UK
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In reply to this post by Aryeh Weiss
Aryeh,
We either run Mark and Find, or Live Data Mode. In the latter case, each Job corresponds to a Z stack at a certain position on the slide, all parameters set up to best conditions. We then add all Jobs to a macro with appropriate time gaps in between consecutive Jobs (Wait/Pause option), and then connect the two ends of the macro and set up the appropriate number of loops. This we do usually for overnight live cell/animal scanning.
With Mark and Find, we choose XYZT mode, visit every location that we want to scan on the slide one-by-one, set up all scan parameters at a given position and THEN (at the end!) we save the position; this way the system will remember all the settings. Then we move to the next position and repeat. As others pointed out already, at the beginning you have to have your motorised stage initalised for both the Live Data Mode and the Mark and Find. In Mark and Find you also have to unclick the "Use same Z stack" option box and must click on the Set Stack (it's called something like this) software button before you move on to the next position.
I hope this helps, Zoltan - Show quoted text -
On Tue, Feb 10, 2009 at 5:46 AM, Aryeh Weiss <[hidden email]> wrote: First -- thanks for the quick responses. We will ask our rep to get us the software upgrade (the system is only a few months old -- I assume that getting the software will not be a problem). -- Zoltan Cseresnyes Facility manager, Imaging Suite University of Cambridge, UK
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In reply to this post by Adrian Smith-6
Adrian,
The Mark and Find still doesn't work in the Live Data Mode, even in version 2. Zoltan On Tue, Feb 10, 2009 at 3:55 AM, Adrian Smith <[hidden email]> wrote:
-- Zoltan Cseresnyes Facility manager, Imaging Suite |
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In reply to this post by Aryeh Weiss
Hi, Zoltan,
The Mark and Find function apparently only works in TCS SP5 mode in 1.8.2. In TCS SP5 mode, the Tile Scan and Mark and Find function buttons are active, but when I switched to Live Data Mode they became greyed out/inactive. It seems to be the nature of the beast. George George Ring, Ph.D. Dept. of Cell and Developmental Biology SUNY Upstate Medical University 750 E. Adams St. Syracuse NY 13210 Tel. (315) 464-8595 FAX (315) 464-8535 email: [hidden email] >>> Zoltan Cseresnyes <[hidden email]> 02/11/09 6:21 PM >>> Adrian, The Mark and Find still doesn't work in the Live Data Mode, even in version 2. Zoltan On Tue, Feb 10, 2009 at 3:55 AM, Adrian Smith <[hidden email] > wrote: > Also, does the Mark & Find function work in Live Data in version 2? > > When I had a quick look at in 1.8.2 it didn't seem like "mark and find" > was available in Live Data mode? (or at least I couldn't see it). > > Regards, > > Adrian Smith > Centenary Institute, Sydney, Australia > > > > On 10/02/2009, at 2:32 PM, Dr. Graham Wright wrote: > > Hello Zoltan & listers, > > You comments suggest that you are a regular user of the Leica Live Data > Mode on the SP5... if so, I was wondering how you handle the data > post-acquisition - to get back to individual time series from each of the > different positions. We tend to use ImageJ and macros we have written to > sort and concatenate the files, but I am interested in how others approach > this. > > Best wishes, > Graham > > ----- Original Message ----- > *From:* Zoltan Cseresnyes <[hidden email]> > *To:* [hidden email] > *Sent:* Tuesday, February 10, 2009 6:36 AM > *Subject:* Re: Leica SP5 question > > Aryeh, > > If you upgrade to LAS AF version 2.0, you'll be able to open a second > window and observe previously acquired data. We very much like the new > version anyway, e.g. we find the Mark and Find and Live Data Mode more > stable than even with the 1.8.2, which was already pretty good. I hope > this helps, > > Zoltan > > > > On Mon, Feb 9, 2009 at 10:31 PM, Aryeh Weiss <[hidden email]> wrote: > >> When acquiring confocal data, we find it useful to have a previous >> experiment (usually an image acquired earlier during that session) open, >> while making the live acquisition. This allows an immediate visual >> comparison of the current result with a previous result. On the Leica >> we do not seem to be able to do this, because apparently only one data >> window can be opened, and if a new scan is done, it replaces the old data in >> that window. >> >> I suppose I can use ImageJ to open the stored data, but i was wondering if >> we are missing something obvious. >> >> --aryeh >> -- >> Aryeh Weiss >> School of Engineering >> Bar Ilan University >> Ramat Gan 52900 Israel >> >> Ph: 972-3-5317638 >> FAX: 972-3-7384050 >> > > > > -- > > Zoltan Cseresnyes > Facility manager, Imaging Suite > University of Cambridge, UK > > > -- Zoltan Cseresnyes Facility manager, Imaging Suite |
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Keith Morris |
Re: Leica SP5 question - Motorised Stage
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In reply to this post by Adrian Smith-6
Hi,
Don't know if the following is off any help at all [or whether you are asking a Leica SP5 stage software specific question], but I this response to similar query a while ago and it may or may not be useful [i.e. I always found it a good idea to visually check what the software is doing physically with the stage, and the following was how I generally did it]: " I know it's going back 15 years since I regularly used a motorised stage* on a standard microscope [other than a PALM micro-dissection system], but I always used my England Finder to ensure that the stage origin 0,0 was always in the same place - assuming the slide & slide holder is fixed/moved to the same point on the stage. The England Finder slide has a complete reference grid etched across its surface. I always aligned the stage origin to a specific point, e.g. a cross or the tip of a letter/number. It's nice to see the 0,0 origin co-ordinate always confirmed as the same place on-screen visually [and you can use it to confirm co-ordinates of objects of interest, although you have to take the specimen slide out and replace it with the England Finder slide]. Our main Magiscan image analyser microscopes had 8 and 16 slide stages so things got complicated. The software allowed us to define any point as the stage origin. Our Magiscan software could even optionally report stage positions in England finder co-ordinates. You can buy slide cover-slips and Mattek dishes with these grids etched on them as well, but only the England Finder has them etched over the entire slide. Once our stage origin 0,0 was sorted out though things were fine. You can use the England Finder to quickly re-locate specimens on different microscopes, once you calculate the conversion factors for different motorised/manual stages. I suppose you could make your own 0,0 reference point using fixed features on any slide, but the entire etched grid is really useful. Although it's easier to set the 0,0 stage origin within the visible slide area, assuming the software allows this [ours always did], you can use the Finder to check the reproducibility of the position of other stage co-ordinates and that the field raster scan is moving correctly. Only downside is that the England Finder is fragile and costs about $250, although both mine came free from a drawer in a lab millennia ago, so you might find one lying about [and twenty five years on mine still aren't chipped]. See one at: http://www.2spi.com/catalog/magnifiers/england-finder-graticule-instructions .html http://www.2spi.com/catalog/magnifiers/england-finder-graticule.shtml I used to calculate things like exact field sizes etc.. for snake and raster scans to ensure the software gets it right. So you need the odd calibration graticule slides as well [1mm/1,000 scale probably, with a 200um one also useful]: http://www.2spi.com/catalog/ltmic/stage-micrometers-graticules.shtml I used to stick a physical rectangular field as 'lines' [Letraset I think] on the screen to align fields in raster scans, but I'm sure software could do it these days. Our systems were all image analysers so checking distances, etc. was easy [I expect your SP5 can display lengths, and it's already been calibrated by Leica]. It's also helpful to see a bit of the area outside the field on-screen to ensure the next field is exactly aligned during raster scans [and to see where things in the field go]. " Hope this is of some help [and I haven't misinterpreted your question]. Plus I'm not familiar with the SP5 stage or its software driver options, & some offer more control than others. But the above worked for me with our setups. Our UCL PALM system set it's own stage origin automatically in hardware/software, but it's motorised stage was pretty unique [and expensive] as it can drive diagonally [for laser cutting] as well in 'straight' steps in X & Y. Regards Keith *Back in the 80s every microscope I bought for my personal research use had a motorised stage, but collective decisions for core facilities have led to most microscopes where I've worked since being supplied with manual XY stages. A mistake in my opinion, particularly as they are discounted when included in the original tender and very expensive to add afterwards on a limited operating budget. --------------------------------------------------------------------------- Dr Keith J. Morris, Molecular Cytogenetics and Microscopy Core, Laboratory 00/069 and 00/070, The Wellcome Trust Centre for Human Genetics, Roosevelt Drive, Oxford OX3 7BN, United Kingdom. Telephone: +44 (0)1865 287568 Email: [hidden email] Web-pages: http://www.well.ox.ac.uk/cytogenetics/ -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Adrian Smith Sent: 10 February 2009 23:01 To: [hidden email] Subject: Re: Leica SP5 question - Motorised Stage On 10/02/2009, at 4:59 PM, Cameron Nowell wrote: > To be able to do XYZT acquisition with multiple fields your > scope needs to be fitted with a motorised stage. If you have that then > you will be fine. While we are talking SP5 and motorised stage.... I've been wondering if anyone has a come up with a good procedure for setting up the calibration of the stage position, ie in the Stage Configuration window there are settings for Tile Scan overlap, origin offset sizes, merge settings that need to be set up in order for tile scans to work. One of the students here got them quite close through a laborious trial-and-error process. Unfortunately, we need to reset them because the stage alignment got messed up at one point (when the stage cable got caught in the opening into the incubation enclosure). Before I go through the process of doing it by trial and error or working out a more systematic way of doing it I was wondering if anyone else had any suggestions as to how best approach it? Regards, Adrian Smith Centenary Institute, Sydney, Australia |
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