Aligning large stacks

[Feinstein, Timothy]

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Hello all,

We have a number of folks who routinely test the limits of our A1-RS system by collecting for example large and high-resolution atlases of clarified brains (at least the 300 um -1 mm depth that we can reach with an inverted scope).  Our coded stage and a software feature in Elements lets us create 3D stacks of very large images stitched together from 16 to 100 individual images per slice.

We have found that the alignment can go slightly off from one slice to the next, screwing with our deconvolution and analysis results.  This is not by itself shocking; these jobs are asking an awful lot of the system.  We have the system pretty well isolated from vibration and air currents and nobody goes near it while collecting.

Elements has an alignment feature built in but it does not produce great results from very large images (e.g. 4096 ^2 and up).  I have tried the registration plugins menu in Fiji and my current favorite, StackReg, can produce solid results.  Rigid body registration seems to do the job most of the time.  On the other hand its progress bar is not very helpful and some alignments can take overnight to all weekend, if they work.

So I thought it would help to throw this out to those of you who do this more often – do you have a favorite workflow for big-job registrations?  Do you have a favorite freeware, Matlab plugin or paid option for these big jobs?  Should I stop stitching the images and let the coded stage handle alignment on its own?

We use a workstation with a decent graphics card and 36 GB of RAM.  We can put in a lot more if that will help.

Any advice on or off list would be appreciated.

Thanks and all the best,


TF

Timothy Feinstein, Ph.D. | Confocal Manager
333 Bostwick Ave., N.E., Grand Rapids, Michigan 49503
Phone: [phone] | Email: [hidden email]<mailto:[hidden email]>

[Johannes Schindelin]

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Hi TF,

On Wed, 2 Apr 2014, Feinstein, Timothy wrote:

> Elements has an alignment feature built in but it does not produce great
> results from very large images (e.g. 4096 ^2 and up).  I have tried the
> registration plugins menu in Fiji and my current favorite, StackReg, can
> produce solid results.  Rigid body registration seems to do the job most
> of the time.  On the other hand its progress bar is not very helpful and
> some alignments can take overnight to all weekend, if they work.
>
> So I thought it would help to throw this out to those of you who do this
> more often – do you have a favorite workflow for big-job registrations?
> Do you have a favorite freeware, Matlab plugin or paid option for these
> big jobs?  Should I stop stitching the images and let the coded stage
> handle alignment on its own?

I had some very good experiences with "Linear Stack Alignment with SIFT"
in Fiji a little over a year ago. My problem was small enough to run
completely in computer RAM, though. And the parameters had to be adjusted
to the images of course.

I also heard that XuvTools (http://www.xuvtools.org/) is supposed to work
quite well, but I never tried (I wanted to, and since it claims to be Open
Source, I wanted to inspect how it does it in true scientific fashion, but
you have to ask for permission to see the source code -- which is actually
not Open source -- so I did not). Your mileage may vary.

Hope this helps!
Johannes

[Straatman, Kees (Dr.)]

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Dear Timothy,

We recently had some problems with stitching using NIS-Elements which were resolved by calibrating the stage.  The info I received from our Nikon product specialist was:

"Go to the Calibration menu and objectives, there is an option on the right to recalibrate objective (ensure the correct objective is selected). In the wizard select the auto option, ensure you have a sample on the stage in focus with good detail. The software will run an autocalibrate and should then stitch correctly."

This solved the problem for us. We also noticed that after the camera was slightly turned by accident the stitching did not work properly anymore and we needed to recalibrate.

Best wishes

Kees


Dr Ir K.R. Straatman
Senior Experimental Officer
Centre for Core Biotechnology Services
University of Leicester
http://www2.le.ac.uk/colleges/medbiopsych/facilities-and-services/cbs/lite/aif




-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Feinstein, Timothy
Sent: 02 April 2014 19:11
To: [hidden email]
Subject: Aligning large stacks

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Hello all,

We have a number of folks who routinely test the limits of our A1-RS system by collecting for example large and high-resolution atlases of clarified brains (at least the 300 um -1 mm depth that we can reach with an inverted scope).  Our coded stage and a software feature in Elements lets us create 3D stacks of very large images stitched together from 16 to 100 individual images per slice.

We have found that the alignment can go slightly off from one slice to the next, screwing with our deconvolution and analysis results.  This is not by itself shocking; these jobs are asking an awful lot of the system.  We have the system pretty well isolated from vibration and air currents and nobody goes near it while collecting.

Elements has an alignment feature built in but it does not produce great results from very large images (e.g. 4096 ^2 and up).  I have tried the registration plugins menu in Fiji and my current favorite, StackReg, can produce solid results.  Rigid body registration seems to do the job most of the time.  On the other hand its progress bar is not very helpful and some alignments can take overnight to all weekend, if they work.

So I thought it would help to throw this out to those of you who do this more often - do you have a favorite workflow for big-job registrations?  Do you have a favorite freeware, Matlab plugin or paid option for these big jobs?  Should I stop stitching the images and let the coded stage handle alignment on its own?

We use a workstation with a decent graphics card and 36 GB of RAM.  We can put in a lot more if that will help.

Any advice on or off list would be appreciated.

Thanks and all the best,


TF

Timothy Feinstein, Ph.D. | Confocal Manager
333 Bostwick Ave., N.E., Grand Rapids, Michigan 49503
Phone: [phone] | Email: [hidden email]<mailto:[hidden email]>

[phil laissue-2]

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Hi Timothy,

like Kees, we're happy with NIS-Elements stitching, overall. Automatic
calibration works best with a high-contrast, continuous sample - tissue
sections (even just HE) work fine, e.g. dog ileum (but not, say, breast
tissue). Stitching overlap between 5-15% is fine. I did run into problems
with too large samples though. A coral I took was ~5k images
(x14*y4*z30*ch3), resulting in around a 5GB nd2 file. NIS-Elements
subsequently crashed trying to put it together, and the file was corrupted.
Luckily, I always save them separately as tifs, and stitched those
together. For ease of handling, I collapsed z and stitched maximum
intensity projections (and extended depth of field for DIC, using the BIG
Lausanne's plugin) together for the big picture, using 3D stitching for
smaller ROIs. MosaicJ works fine, as does Stephan Preibisch's.
StackReg (BIG Lausanne) often works ok (I only stick with Translation or
rigid body, never scaled or affine for obvious reasons). Especially useful
is a version using StackReg, called MultiStackReg or MultiStackRegFix (Brad
Busse/Ping Fu & Jennifer Staab, resp.), allow saving the transformation
matrix for one channel and aligning the next using those parameters. I'd
like to do the same for the SIFT-based registration plugin (Stephan
Saalfeld), which mostly works really well (parameters sometimes need
adjusting) and often gets the job done where StackReg fails.

Kind regards

Philippe

_____________________________________
Philippe Laissue, PhD, Bioimaging Manager
School of Biological Sciences, Room 4.17
University of Essex, Colchester CO4 3SQ, UK
(0044) 01206 872246 / (0044) 07842 676 456
[hidden email]
privatewww.essex.ac.uk/~plaissue <http://privatewww.essex.ac.uk/%7Eplaissue>




_____________________________________
Philippe Laissue, PhD, Bioimaging Manager
School of Biological Sciences, Room 4.17
University of Essex, Colchester CO4 3SQ, UK
(0044) 01206 872246 / (0044) 07842 676 456
[hidden email]
privatewww.essex.ac.uk/~plaissue


On 3 April 2014 09:05, Straatman, Kees (Dr.) <[hidden email]> wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> Post images on http://www.imgur.com and include the link in your posting.
> *****
>
> Dear Timothy,
>
> We recently had some problems with stitching using NIS-Elements which were
> resolved by calibrating the stage.  The info I received from our Nikon
> product specialist was:
>
> "Go to the Calibration menu and objectives, there is an option on the
> right to recalibrate objective (ensure the correct objective is selected).
> In the wizard select the auto option, ensure you have a sample on the stage
> in focus with good detail. The software will run an autocalibrate and
> should then stitch correctly."
>
> This solved the problem for us. We also noticed that after the camera was
> slightly turned by accident the stitching did not work properly anymore and
> we needed to recalibrate.
>
> Best wishes
>
> Kees
>
>
> Dr Ir K.R. Straatman
> Senior Experimental Officer
> Centre for Core Biotechnology Services
> University of Leicester
>
> http://www2.le.ac.uk/colleges/medbiopsych/facilities-and-services/cbs/lite/aif
>
>
>
>
> -----Original Message-----
> From: Confocal Microscopy List [mailto:[hidden email]]
> On Behalf Of Feinstein, Timothy
> Sent: 02 April 2014 19:11
> To: [hidden email]
> Subject: Aligning large stacks
>
> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> Post images on http://www.imgur.com and include the link in your posting.
> *****
>
> Hello all,
>
> We have a number of folks who routinely test the limits of our A1-RS
> system by collecting for example large and high-resolution atlases of
> clarified brains (at least the 300 um -1 mm depth that we can reach with an
> inverted scope).  Our coded stage and a software feature in Elements lets
> us create 3D stacks of very large images stitched together from 16 to 100
> individual images per slice.
>
> We have found that the alignment can go slightly off from one slice to the
> next, screwing with our deconvolution and analysis results.  This is not by
> itself shocking; these jobs are asking an awful lot of the system.  We have
> the system pretty well isolated from vibration and air currents and nobody
> goes near it while collecting.
>
> Elements has an alignment feature built in but it does not produce great
> results from very large images (e.g. 4096 ^2 and up).  I have tried the
> registration plugins menu in Fiji and my current favorite, StackReg, can
> produce solid results.  Rigid body registration seems to do the job most of
> the time.  On the other hand its progress bar is not very helpful and some
> alignments can take overnight to all weekend, if they work.
>
> So I thought it would help to throw this out to those of you who do this
> more often - do you have a favorite workflow for big-job registrations?  Do
> you have a favorite freeware, Matlab plugin or paid option for these big
> jobs?  Should I stop stitching the images and let the coded stage handle
> alignment on its own?
>
> We use a workstation with a decent graphics card and 36 GB of RAM.  We can
> put in a lot more if that will help.
>
> Any advice on or off list would be appreciated.
>
> Thanks and all the best,
>
>
> TF
>
> Timothy Feinstein, Ph.D. | Confocal Manager
> 333 Bostwick Ave., N.E., Grand Rapids, Michigan 49503
> Phone: [phone] | Email: [hidden email]<mailto:
> [hidden email]>
>

[Feinstein, Timothy]

*****
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*****

Thanks again to everyone who replied on and off list.  We calibrated the
objectives as Kees recommended and still found some misalignment.  We next
tried the same with stitching turned off and the alignment was perfect.
Blurring of fine details was a little worse at image borders but not by
very much.  It turns out that the coded stage really is very good at
maintaining a position when imaging 100+ tiled images repeatedly in a
large number on z sections.

All the best,


TF

Timothy Feinstein, Ph.D. | Confocal Manager
333 Bostwick Ave., N.E., Grand Rapids, Michigan 49503
Phone: [phone] | Email: [hidden email]







On 4/3/14, 9:36 AM, "phil laissue" <[hidden email]> wrote:

>*****
>To join, leave or search the confocal microscopy listserv, go to:
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>GkAd6xg&u=http%3a%2f%2flists%2eumn%2eedu%2fcgi-bin%2fwa%3fA0%3dconfocalmic
>roscopy
>Post images on
>http://scanmail.trustwave.com/?c=129&d=3Ou907qXF6gXlgdAfyFZpLpgH38HKJr2HDh
>Hwg0ulg&u=http%3a%2f%2fwww%2eimgur%2ecom and include the link in your
>posting.
>*****
>
>Hi Timothy,
>
>like Kees, we're happy with NIS-Elements stitching, overall. Automatic
>calibration works best with a high-contrast, continuous sample - tissue
>sections (even just HE) work fine, e.g. dog ileum (but not, say, breast
>tissue). Stitching overlap between 5-15% is fine. I did run into problems
>with too large samples though. A coral I took was ~5k images
>(x14*y4*z30*ch3), resulting in around a 5GB nd2 file. NIS-Elements
>subsequently crashed trying to put it together, and the file was
>corrupted.
>Luckily, I always save them separately as tifs, and stitched those
>together. For ease of handling, I collapsed z and stitched maximum
>intensity projections (and extended depth of field for DIC, using the BIG
>Lausanne's plugin) together for the big picture, using 3D stitching for
>smaller ROIs. MosaicJ works fine, as does Stephan Preibisch's.
>StackReg (BIG Lausanne) often works ok (I only stick with Translation or
>rigid body, never scaled or affine for obvious reasons). Especially useful
>is a version using StackReg, called MultiStackReg or MultiStackRegFix
>(Brad
>Busse/Ping Fu & Jennifer Staab, resp.), allow saving the transformation
>matrix for one channel and aligning the next using those parameters. I'd
>like to do the same for the SIFT-based registration plugin (Stephan
>Saalfeld), which mostly works really well (parameters sometimes need
>adjusting) and often gets the job done where StackReg fails.
>
>Kind regards
>
>Philippe
>
>_____________________________________
>Philippe Laissue, PhD, Bioimaging Manager
>School of Biological Sciences, Room 4.17
>University of Essex, Colchester CO4 3SQ, UK
>(0044) 01206 872246 / (0044) 07842 676 456
>[hidden email]
>http://scanmail.trustwave.com/?c=129&d=3eu904ma0h6a25H5kaVFITC-PSzasnHZIGe
>V2W8npA&u=http%3a%2f%2fprivatewww%2eessex%2eac%2euk%2f%7eplaissue
><http://scanmail.trustwave.com/?c=129&d=3eu904ma0h6a25H5kaVFITC-PSzasnHZID
>3BiWh78g&u=http%3a%2f%2fprivatewww%2eessex%2eac%2euk%2f%257Eplaissue>
>
>
>
>
>_____________________________________
>Philippe Laissue, PhD, Bioimaging Manager
>School of Biological Sciences, Room 4.17
>University of Essex, Colchester CO4 3SQ, UK
>(0044) 01206 872246 / (0044) 07842 676 456
>[hidden email]
>http://scanmail.trustwave.com/?c=129&d=3eu904ma0h6a25H5kaVFITC-PSzasnHZIGe
>V2W8npA&u=http%3a%2f%2fprivatewww%2eessex%2eac%2euk%2f%7eplaissue
>
>
>On 3 April 2014 09:05, Straatman, Kees (Dr.) <[hidden email]> wrote:
>
>> *****
>> To join, leave or search the confocal microscopy listserv, go to:
>>
>>http://scanmail.trustwave.com/?c=129&d=3eu904ma0h6a25H5kaVFITC-PSzasnHZID
>>LC32Z3pg&u=http%3a%2f%2flists%2eumn%2eedu%2fcgi-bin%2fwa%3fA0%3dconfocalm
>>icroscopy
>> Post images on
>>http://scanmail.trustwave.com/?c=129&d=3eu904ma0h6a25H5kaVFITC-PSzasnHZIG
>>fDjWwj9g&u=http%3a%2f%2fwww%2eimgur%2ecom and include the link in your
>>posting.
>> *****
>>
>> Dear Timothy,
>>
>> We recently had some problems with stitching using NIS-Elements which
>>were
>> resolved by calibrating the stage.  The info I received from our Nikon
>> product specialist was:
>>
>> "Go to the Calibration menu and objectives, there is an option on the
>> right to recalibrate objective (ensure the correct objective is
>>selected).
>> In the wizard select the auto option, ensure you have a sample on the
>>stage
>> in focus with good detail. The software will run an autocalibrate and
>> should then stitch correctly."
>>
>> This solved the problem for us. We also noticed that after the camera
>>was
>> slightly turned by accident the stitching did not work properly anymore
>>and
>> we needed to recalibrate.
>>
>> Best wishes
>>
>> Kees
>>
>>
>> Dr Ir K.R. Straatman
>> Senior Experimental Officer
>> Centre for Core Biotechnology Services
>> University of Leicester
>>
>>
>>http://scanmail.trustwave.com/?c=129&d=3eu904ma0h6a25H5kaVFITC-PSzasnHZID
>>aV3z8n9A&u=http%3a%2f%2fwww2%2ele%2eac%2euk%2fcolleges%2fmedbiopsych%2ffa
>>cilities-and-services%2fcbs%2flite%2faif
>>
>>
>>
>>
>> -----Original Message-----
>> From: Confocal Microscopy List [mailto:[hidden email]]
>> On Behalf Of Feinstein, Timothy
>> Sent: 02 April 2014 19:11
>> To: [hidden email]
>> Subject: Aligning large stacks
>>
>> *****
>> To join, leave or search the confocal microscopy listserv, go to:
>>
>>http://scanmail.trustwave.com/?c=129&d=3eu904ma0h6a25H5kaVFITC-PSzasnHZID
>>LC32Z3pg&u=http%3a%2f%2flists%2eumn%2eedu%2fcgi-bin%2fwa%3fA0%3dconfocalm
>>icroscopy
>> Post images on
>>http://scanmail.trustwave.com/?c=129&d=3eu904ma0h6a25H5kaVFITC-PSzasnHZIG
>>fDjWwj9g&u=http%3a%2f%2fwww%2eimgur%2ecom and include the link in your
>>posting.
>> *****
>>
>> Hello all,
>>
>> We have a number of folks who routinely test the limits of our A1-RS
>> system by collecting for example large and high-resolution atlases of
>> clarified brains (at least the 300 um -1 mm depth that we can reach
>>with an
>> inverted scope).  Our coded stage and a software feature in Elements
>>lets
>> us create 3D stacks of very large images stitched together from 16 to
>>100
>> individual images per slice.
>>
>> We have found that the alignment can go slightly off from one slice to
>>the
>> next, screwing with our deconvolution and analysis results.  This is
>>not by
>> itself shocking; these jobs are asking an awful lot of the system.  We
>>have
>> the system pretty well isolated from vibration and air currents and
>>nobody
>> goes near it while collecting.
>>
>> Elements has an alignment feature built in but it does not produce great
>> results from very large images (e.g. 4096 ^2 and up).  I have tried the
>> registration plugins menu in Fiji and my current favorite, StackReg, can
>> produce solid results.  Rigid body registration seems to do the job
>>most of
>> the time.  On the other hand its progress bar is not very helpful and
>>some
>> alignments can take overnight to all weekend, if they work.
>>
>> So I thought it would help to throw this out to those of you who do this
>> more often - do you have a favorite workflow for big-job registrations?
>> Do
>> you have a favorite freeware, Matlab plugin or paid option for these big
>> jobs?  Should I stop stitching the images and let the coded stage handle
>> alignment on its own?
>>
>> We use a workstation with a decent graphics card and 36 GB of RAM.  We
>>can
>> put in a lot more if that will help.
>>
>> Any advice on or off list would be appreciated.
>>
>> Thanks and all the best,
>>
>>
>> TF
>>
>> Timothy Feinstein, Ph.D. | Confocal Manager
>> 333 Bostwick Ave., N.E., Grand Rapids, Michigan 49503
>> Phone: [phone] | Email: [hidden email]<mailto:
>> [hidden email]>
>>

[Craig Brideau]

*****
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Post images on http://www.imgur.com and include the link in your posting.
*****

Good stages have proper gear backlash correction and the like (takes up the
slop between the teeth in the gears), so with a decent motion controller
and encoding you can expect good accuracy. Some stages are starting to
feature linear motor drive systems which even eliminate the gear backlash
issue, so we can only expect things to get even better!

Craig


On Sun, Apr 6, 2014 at 1:05 PM, Feinstein, Timothy <
[hidden email]> wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> Post images on http://www.imgur.com and include the link in your posting.
> *****
>
> Thanks again to everyone who replied on and off list.  We calibrated the
> objectives as Kees recommended and still found some misalignment.  We next
> tried the same with stitching turned off and the alignment was perfect.
> Blurring of fine details was a little worse at image borders but not by
> very much.  It turns out that the coded stage really is very good at
> maintaining a position when imaging 100+ tiled images repeatedly in a
> large number on z sections.
>
> All the best,
>
>
> TF
>
> Timothy Feinstein, Ph.D. | Confocal Manager
> 333 Bostwick Ave., N.E., Grand Rapids, Michigan 49503
> Phone: [phone] | Email: [hidden email]
>
>
>
>
>
>
>
> On 4/3/14, 9:36 AM, "phil laissue" <[hidden email]> wrote:
>
> >*****
> >To join, leave or search the confocal microscopy listserv, go to:
> >
> http://scanmail.trustwave.com/?c=129&d=3Ou907qXF6gXlgdAfyFZpLpgH38HKJr2HG1
> >GkAd6xg&u=http%3a%2f%2flists%2eumn%2eedu%2fcgi-bin%2fwa%3fA0%3dconfocalmic
> >roscopy
> >Post images on
> >
> http://scanmail.trustwave.com/?c=129&d=3Ou907qXF6gXlgdAfyFZpLpgH38HKJr2HDh
> >Hwg0ulg&u=http%3a%2f%2fwww%2eimgur%2ecom and include the link in your
> >posting.
> >*****
> >
> >Hi Timothy,
> >
> >like Kees, we're happy with NIS-Elements stitching, overall. Automatic
> >calibration works best with a high-contrast, continuous sample - tissue
> >sections (even just HE) work fine, e.g. dog ileum (but not, say, breast
> >tissue). Stitching overlap between 5-15% is fine. I did run into problems
> >with too large samples though. A coral I took was ~5k images
> >(x14*y4*z30*ch3), resulting in around a 5GB nd2 file. NIS-Elements
> >subsequently crashed trying to put it together, and the file was
> >corrupted.
> >Luckily, I always save them separately as tifs, and stitched those
> >together. For ease of handling, I collapsed z and stitched maximum
> >intensity projections (and extended depth of field for DIC, using the BIG
> >Lausanne's plugin) together for the big picture, using 3D stitching for
> >smaller ROIs. MosaicJ works fine, as does Stephan Preibisch's.
> >StackReg (BIG Lausanne) often works ok (I only stick with Translation or
> >rigid body, never scaled or affine for obvious reasons). Especially useful
> >is a version using StackReg, called MultiStackReg or MultiStackRegFix
> >(Brad
> >Busse/Ping Fu & Jennifer Staab, resp.), allow saving the transformation
> >matrix for one channel and aligning the next using those parameters. I'd
> >like to do the same for the SIFT-based registration plugin (Stephan
> >Saalfeld), which mostly works really well (parameters sometimes need
> >adjusting) and often gets the job done where StackReg fails.
> >
> >Kind regards
> >
> >Philippe
> >
> >_____________________________________
> >Philippe Laissue, PhD, Bioimaging Manager
> >School of Biological Sciences, Room 4.17
> >University of Essex, Colchester CO4 3SQ, UK
> >(0044) 01206 872246 / (0044) 07842 676 456
> >[hidden email]
> >
> http://scanmail.trustwave.com/?c=129&d=3eu904ma0h6a25H5kaVFITC-PSzasnHZIGe
> >V2W8npA&u=http%3a%2f%2fprivatewww%2eessex%2eac%2euk%2f%7eplaissue
> ><
> http://scanmail.trustwave.com/?c=129&d=3eu904ma0h6a25H5kaVFITC-PSzasnHZID
> >3BiWh78g&u=http%3a%2f%2fprivatewww%2eessex%2eac%2euk%2f%257Eplaissue>
> >
> >
> >
> >
> >_____________________________________
> >Philippe Laissue, PhD, Bioimaging Manager
> >School of Biological Sciences, Room 4.17
> >University of Essex, Colchester CO4 3SQ, UK
> >(0044) 01206 872246 / (0044) 07842 676 456
> >[hidden email]
> >
> http://scanmail.trustwave.com/?c=129&d=3eu904ma0h6a25H5kaVFITC-PSzasnHZIGe
> >V2W8npA&u=http%3a%2f%2fprivatewww%2eessex%2eac%2euk%2f%7eplaissue
> >
> >
> >On 3 April 2014 09:05, Straatman, Kees (Dr.) <[hidden email]>
> wrote:
> >
> >> *****
> >> To join, leave or search the confocal microscopy listserv, go to:
> >>
> >>
> http://scanmail.trustwave.com/?c=129&d=3eu904ma0h6a25H5kaVFITC-PSzasnHZID
> >>LC32Z3pg&u=http%3a%2f%2flists%2eumn%2eedu%2fcgi-bin%2fwa%3fA0%3dconfocalm
> >>icroscopy
> >> Post images on
> >>
> http://scanmail.trustwave.com/?c=129&d=3eu904ma0h6a25H5kaVFITC-PSzasnHZIG
> >>fDjWwj9g&u=http%3a%2f%2fwww%2eimgur%2ecom and include the link in your
> >>posting.
> >> *****
> >>
> >> Dear Timothy,
> >>
> >> We recently had some problems with stitching using NIS-Elements which
> >>were
> >> resolved by calibrating the stage.  The info I received from our Nikon
> >> product specialist was:
> >>
> >> "Go to the Calibration menu and objectives, there is an option on the
> >> right to recalibrate objective (ensure the correct objective is
> >>selected).
> >> In the wizard select the auto option, ensure you have a sample on the
> >>stage
> >> in focus with good detail. The software will run an autocalibrate and
> >> should then stitch correctly."
> >>
> >> This solved the problem for us. We also noticed that after the camera
> >>was
> >> slightly turned by accident the stitching did not work properly anymore
> >>and
> >> we needed to recalibrate.
> >>
> >> Best wishes
> >>
> >> Kees
> >>
> >>
> >> Dr Ir K.R. Straatman
> >> Senior Experimental Officer
> >> Centre for Core Biotechnology Services
> >> University of Leicester
> >>
> >>
> >>
> http://scanmail.trustwave.com/?c=129&d=3eu904ma0h6a25H5kaVFITC-PSzasnHZID
> >>aV3z8n9A&u=http%3a%2f%2fwww2%2ele%2eac%2euk%2fcolleges%2fmedbiopsych%2ffa
> >>cilities-and-services%2fcbs%2flite%2faif
> >>
> >>
> >>
> >>
> >> -----Original Message-----
> >> From: Confocal Microscopy List [mailto:[hidden email]
> ]
> >> On Behalf Of Feinstein, Timothy
> >> Sent: 02 April 2014 19:11
> >> To: [hidden email]
> >> Subject: Aligning large stacks
> >>
> >> *****
> >> To join, leave or search the confocal microscopy listserv, go to:
> >>
> >>
> http://scanmail.trustwave.com/?c=129&d=3eu904ma0h6a25H5kaVFITC-PSzasnHZID
> >>LC32Z3pg&u=http%3a%2f%2flists%2eumn%2eedu%2fcgi-bin%2fwa%3fA0%3dconfocalm
> >>icroscopy
> >> Post images on
> >>
> http://scanmail.trustwave.com/?c=129&d=3eu904ma0h6a25H5kaVFITC-PSzasnHZIG
> >>fDjWwj9g&u=http%3a%2f%2fwww%2eimgur%2ecom and include the link in your
> >>posting.
> >> *****
> >>
> >> Hello all,
> >>
> >> We have a number of folks who routinely test the limits of our A1-RS
> >> system by collecting for example large and high-resolution atlases of
> >> clarified brains (at least the 300 um -1 mm depth that we can reach
> >>with an
> >> inverted scope).  Our coded stage and a software feature in Elements
> >>lets
> >> us create 3D stacks of very large images stitched together from 16 to
> >>100
> >> individual images per slice.
> >>
> >> We have found that the alignment can go slightly off from one slice to
> >>the
> >> next, screwing with our deconvolution and analysis results.  This is
> >>not by
> >> itself shocking; these jobs are asking an awful lot of the system.  We
> >>have
> >> the system pretty well isolated from vibration and air currents and
> >>nobody
> >> goes near it while collecting.
> >>
> >> Elements has an alignment feature built in but it does not produce great
> >> results from very large images (e.g. 4096 ^2 and up).  I have tried the
> >> registration plugins menu in Fiji and my current favorite, StackReg, can
> >> produce solid results.  Rigid body registration seems to do the job
> >>most of
> >> the time.  On the other hand its progress bar is not very helpful and
> >>some
> >> alignments can take overnight to all weekend, if they work.
> >>
> >> So I thought it would help to throw this out to those of you who do this
> >> more often - do you have a favorite workflow for big-job registrations?
> >> Do
> >> you have a favorite freeware, Matlab plugin or paid option for these big
> >> jobs?  Should I stop stitching the images and let the coded stage handle
> >> alignment on its own?
> >>
> >> We use a workstation with a decent graphics card and 36 GB of RAM.  We
> >>can
> >> put in a lot more if that will help.
> >>
> >> Any advice on or off list would be appreciated.
> >>
> >> Thanks and all the best,
> >>
> >>
> >> TF
> >>
> >> Timothy Feinstein, Ph.D. | Confocal Manager
> >> 333 Bostwick Ave., N.E., Grand Rapids, Michigan 49503
> >> Phone: [phone] | Email: [hidden email]<mailto:
> >> [hidden email]>
> >>
>

[Renato A. Mortara]

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

Dear all

A friend has just purchased  Nikon  DS-Ri1 camera with a DS-U3 controller,
and he is looking for an open source software to control it. Apparently
Micro-Manager does not communicate with the camerea.

Any ideas ?

Thanks !

Renato


Dr. Renato Arruda Mortara
Disciplina de Parasitologia
Escola Paulista de Medicina - UNIFESP
Rua Botucatu, 862 6o. andar
04023-062
São Paulo SP
Brasil
Fone: 11 5579-8306
VOIP: 5576-4848 R: 2928
[hidden email]
www.ecb.epm.br/~ramortara
 

[Kurt Thorn]

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

On 5/23/2014 9:49 AM, Renato Mortara wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> Post images on http://www.imgur.com and include the link in your posting.
> *****
>
> Dear all
>
> A friend has just purchased  Nikon  DS-Ri1 camera with a DS-U3 controller,
> and he is looking for an open source software to control it. Apparently
> Micro-Manager does not communicate with the camerea.
>
> Any ideas ?

As far as I know the Nikon cameras are only controllable with NIS-Elements.

Kurt

>
> Thanks !
>
> Renato
>
>
> Dr. Renato Arruda Mortara
> Disciplina de Parasitologia
> Escola Paulista de Medicina - UNIFESP
> Rua Botucatu, 862 6o. andar
> 04023-062
> São Paulo SP
> Brasil
> Fone: 11 5579-8306
> VOIP: 5576-4848 R: 2928
> [hidden email]
> www.ecb.epm.br/~ramortara
>
>
>

--
Kurt Thorn
Director, Nikon Imaging Center
http://nic.ucsf.edu/blog/