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Hi All,
Thanks for all your messages. Louis described much better than I did what we plan to do "In fact you want to add multiple Z-series together in order to form one huge series"!
We have Volocity (and obviously imageJ) to visualise 3D data and I believe we can add z-series together, on top of each other. My worry is we will have issues aligning structures in one z-stack, from one tissue section, with the same structures in the next z-stack/tissue section.
I'll give it a go using the suggestions made and come back to you if I get stuck!
Thanks again,
Em
-----Original Message-----
From: Confocal Microscopy List [mailto:[log in to unmask]] On Behalf Of CONFOCALMICROSCOPY automatic digest system
Sent: 19 November 2011 06:05
To: [log in to unmask]
Subject: CONFOCALMICROSCOPY Digest - 17 Nov 2011 to 18 Nov 2011 (#2011-88)
There are 12 messages totalling 698 lines in this issue.
Topics of the day:
1. 3D reconstruction of multiple, fluorescently labelled, sections (3)
2. RE 3D reconstruction of multiple, fluorescently labelled, sections (3)
3. Olympus Corp in financial trouble? (2)
4. (lots) more on Olympus
5. Cell viability assay
6. Fast two-photon imaging: Constant fraction discriminator and variable
delay for Laser pulse-synch
7. Olympus Corp in financial trouble? We certainly hope not!
----------------------------------------------------------------------
Date: Fri, 18 Nov 2011 08:39:57 -0600
From: Emma King <[log in to unmask]>
Subject: 3D reconstruction of multiple, fluorescently labelled, sections
*****
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We would like to take serial sections (20 microns thick) of rat spinal=20=
cord (up to 1cm long), label them with a bright and specific fluorescent=20=
label, acquire images (we can use either a widefield or confocal=20
microscope) and then reconstruct the data in to one, 3D structure.=20
We have the sectioning, staining and acquisition parameters/options=20
under control, but don't have any software capable of reconstructing=20
the resultant images. Does anyone have any suggestions as to what=20
software to use and where to get it from?=20
Any help much appreciated.=20
Cheers,=20
Emma=20
Advanced Microscopy Unit=20
School of Biomedical Sciences=20
Univeristy of Nottingham
------------------------------
Date: Fri, 18 Nov 2011 09:51:43 -0500
From: [log in to unmask]
Subject: RE 3D reconstruction of multiple, fluorescently labelled, sections
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Hi Emma,
In fact you want to add mutltiple Z-series together in order to form one=20
huge serie. First, you will need a good computer and 2nd, I have done it=20
using Huygens software (SVI).=20
Also, I am pretty sure that there is a plugin In" Image J" that can do=20
that. Do not forget to do that with a good computer!!!!
Best regards,
Louis
Louis Villeneuve
Associ=E9 de recherche - Microscopie confocale
Institut de Cardiologie de Montreal- Centre de recherche
5000 est B=E9langer
Montreal (Qc), Canada
H1T 3C8
514-376-3330 ext 3511=20
514-376-1355 (fax)
Emma King <[log in to unmask]>@LISTS.UMN.EDU=20
Envoy=E9 par : Confocal Microscopy List <[log in to unmask]>
2011-11-18 09:39
Veuillez r=E9pondre =E0
Confocal Microscopy List <[log in to unmask]>
A
[log in to unmask]
cc
Objet
3D reconstruction of multiple, fluorescently labelled, sections
*****
To join, leave or search the confocal microscopy listserv, go to:
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*****
We would like to take serial sections (20 microns thick) of rat spinal=20
cord (up to 1cm long), label them with a bright and specific fluorescent=20
label, acquire images (we can use either a widefield or confocal=20
microscope) and then reconstruct the data in to one, 3D structure.=20
We have the sectioning, staining and acquisition parameters/options=20
under control, but don't have any software capable of reconstructing=20
the resultant images. Does anyone have any suggestions as to what=20
software to use and where to get it from?=20
Any help much appreciated.=20
Cheers,=20
Emma=20
Advanced Microscopy Unit=20
School of Biomedical Sciences=20
Univeristy of Nottingham
------------------------------
Date: Fri, 18 Nov 2011 09:56:23 -0500
From: Louis Kerr <[log in to unmask]>
Subject: Re: 3D reconstruction of multiple, fluorescently labelled, sections
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Hi Emma,
Have you seen the work from Stephen Smith's Lab at Stanford with array tomography? This is a link to a Cold Spring Harbor Protocol.
http :// cshprotocols . cshlp .org/content/2010/11/ pdb .top89.full
Good luck,
Louie
----- Original Message -----
From: "Emma King" < emma [log in to unmask]>
To: CONFOCALMICROSCOPY @LISTS. UMN . EDU
Sent: Friday, November 18, 2011 9:39:57 AM
Subject: 3D reconstruction of multiple, fluorescently labelled , sections
*****
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http ://lists. umn . edu /cgi-bin/ wa ?A0= confocalmicroscopy
*****
We would like to take serial sections (20 microns thick) of rat spinal
cord (up to 1cm long), label them with a bright and specific fluorescent
label, acquire images (we can use either a widefield or confocal
microscope) and then reconstruct the data in to one, 3D structure.
We have the sectioning, staining and acquisition parameters/options
under control, but don't have any software capable of reconstructing
the resultant images. Does anyone have any suggestions as to what
software to use and where to get it from?
Any help much appreciated.
Cheers,
Emma
Advanced Microscopy Unit
School of Biomedical Sciences
Univeristy of Nottingham
--
Louis Kerr
lkerr @ mbl . edu
Research and Education Support Coordinator
Marine Biological Laboratory
7 MBL Street
Woods Hole, MA 02543
508-289-7273
508-540-6902 (FAX)
508-292-0289 (Cell phone)
VISIT OUR WEB SITES:
www . mbl . edu
www .courses. mbl . edu
------------------------------
Date: Fri, 18 Nov 2011 16:51:30 +0100
From: Tineke Vendrig <[log in to unmask]>
Subject: Re: 3D reconstruction of multiple, fluorescently labelled, sections
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We use the Andor software for our 3D movies, works nice and good!
Tineke Vendrig
--
Tineke Vendrig, ing
technical engineer optical microscopy
Delft University of Technology
Bionano Science
Kavli Institute of Nanoscience
Lorentzweg 1
2628LJ Delft
room F185
Tel: +31 15 2789299
Fax:+31 15 2781202
email: [log in to unmask]
mobile phone: +31 624341412
2011/11/18 Emma King <[log in to unmask]>
> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> *****
>
> We would like to take serial sections (20 microns thick) of rat spinal
> cord (up to 1cm long), label them with a bright and specific fluorescent
> label, acquire images (we can use either a widefield or confocal
> microscope) and then reconstruct the data in to one, 3D structure.
>
> We have the sectioning, staining and acquisition parameters/options
> under control, but don't have any software capable of reconstructing
> the resultant images. Does anyone have any suggestions as to what
> software to use and where to get it from?
>
> Any help much appreciated.
>
> Cheers,
> Emma
>
> Advanced Microscopy Unit
> School of Biomedical Sciences
> Univeristy of Nottingham
>
------------------------------
Date: Fri, 18 Nov 2011 12:28:12 -0500
From: "Cammer, Michael" <[log in to unmask]>
Subject: Olympus Corp in financial trouble?
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"Japanese officials say that at least $4.9 billion is unaccounted for in a =
financial scandal at Olympus<http://topics.nytimes.com/top/news/business/co=
mpanies/olympus_corporation/index.html?inline=3Dnyt-org> and are investigat=
ing whether much of that money went to companies with links to organized cr=
ime."
http://www.nytimes.com/2011/11/18/business/global/japanese-police-investiga=
te-olympus.html?_r=3D1&scp=3D3&sq=3Dolympus&st=3Dcse
________________________________________________________
Michael Cammer, Assistant Research Scientist
Skirball Institute of Biomolecular Medicine
Lab: (212) 263-3208 Cell: (914) 309-3270
</PRE>
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<PRE>
------------------------------
Date: Fri, 18 Nov 2011 12:18:55 -0600
From: Johannes Schindelin <[log in to unmask]>
Subject: Re: RE 3D reconstruction of multiple, fluorescently labelled, sections
This message is in MIME format. The first part should be readable text,
while the remaining parts are likely unreadable without MIME-aware tools.
--Boundary_(ID_BmV8ySyi3ZWYFpVvpZWp+A)
Content-type: TEXT/PLAIN; charset=ISO-8859-15
Content-transfer-encoding: 8BIT
*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
*****
On Fri, 18 Nov 2011, [log in to unmask] wrote:
> Emma King <[log in to unmask]>@LISTS.UMN.EDU
> Envoyé par : Confocal Microscopy List <[log in to unmask]>
> 2011-11-18 09:39
>
> > We would like to take serial sections (20 microns thick) of rat spinal
> > cord (up to 1cm long), label them with a bright and specific
> > fluorescent label, acquire images (we can use either a widefield or
> > confocal microscope) and then reconstruct the data in to one, 3D
> > structure.
>
> > We have the sectioning, staining and acquisition parameters/options
> > under control, but don't have any software capable of reconstructing
> > the resultant images. Does anyone have any suggestions as to what
> > software to use and where to get it from?
>
> In fact you want to add mutltiple Z-series together in order to form one
> huge serie. First, you will need a good computer and 2nd, I have done
> it using Huygens software (SVI).
>
> Also, I am pretty sure that there is a plugin In" Image J" that can do
> that. Do not forget to do that with a good computer!!!!
If all you want to do is a volume rendering, there are indeed 2 ImageJ
plugins to do the job: Volume Viewer (non-accelerated) and 3D Viewer (this
one uses your graphics adapter's accelerated 3D rendering).
However, if you need to have surface models, you might want to look into
the TrakEM2 plugin. It is very powerful, therefore it takes some training
before one can exploit its capabilities in full.
If you do not want to bother to find all the bits and pieces required for
these plugins to run, please feel free to download Fiji (http://fiji.sc/)
which bundles ImageJ with a lot of plugins (including the 3 mentioned
above).
Ciao,
Johannes
--Boundary_(ID_BmV8ySyi3ZWYFpVvpZWp+A)--
------------------------------
Date: Fri, 18 Nov 2011 11:02:21 -0800
From: "Armstrong, Brian" <[log in to unmask]>
Subject: Re: RE 3D reconstruction of multiple, fluorescently labelled, sections
*****
To join, leave or search the confocal microscopy listserv, go to:
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*****
You could try Reconstruct from NIH Human Brain project =
(http://synapses.clm.utexas.edu/tools/reconstruct/reconstruct.stm ).
In my opinion, Amira, Imaris, and Volocity, are probably best at 3D =
visualization tools.=20
Cheers,
Brian D Armstrong PhD
Assistant Research Professor
Light Microscopy Core=20
Beckman Research Institute
City of Hope
Dept of Neuroscience
1450 E Duarte Rd
Duarte, CA 91010
626-256-4673 x62872
http://www.cityofhope.org/research/support/Light-Microscopy-Digital-Imaging=
/Pages/default.aspx
-----Original Message-----
=46rom: Confocal Microscopy List [mailto:[log in to unmask]] =
=
On Behalf Of Johannes Schindelin
Sent: Friday, November 18, 2011 10:19 AM
To: [log in to unmask]
Subject: Re: RE 3D reconstruction of multiple, fluorescently labelled, =
sections
*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa=3FA0=3Dconfocalmicroscopy
*****
On Fri, 18 Nov 2011, [log in to unmask] wrote:
> Emma King <[log in to unmask]>@LISTS.UMN.EDU=20
> Envoy=E9 par : Confocal Microscopy List <[log in to unmask]>
> 2011-11-18 09:39
>
> > We would like to take serial sections (20 microns thick) of rat spinal
> > cord (up to 1cm long), label them with a bright and specific
> > fluorescent label, acquire images (we can use either a widefield or
> > confocal microscope) and then reconstruct the data in to one, 3D
> > structure.=20
>=20
> > We have the sectioning, staining and acquisition parameters/options
> > under control, but don't have any software capable of reconstructing
> > the resultant images. Does anyone have any suggestions as to what
> > software to use and where to get it from=3F=20
>=20
> In fact you want to add mutltiple Z-series together in order to form one
> huge serie. First, you will need a good computer and 2nd, I have done
> it using Huygens software (SVI).=20
>
> Also, I am pretty sure that there is a plugin In" Image J" that can do=20
> that. Do not forget to do that with a good computer!!!!
If all you want to do is a volume rendering, there are indeed 2 ImageJ
plugins to do the job: Volume Viewer (non-accelerated) and 3D Viewer (this
one uses your graphics adapter's accelerated 3D rendering).
However, if you need to have surface models, you might want to look into
the TrakEM2 plugin. It is very powerful, therefore it takes some training
before one can exploit its capabilities in full.
If you do not want to bother to find all the bits and pieces required for
these plugins to run, please feel free to download Fiji (http://fiji.sc/)
which bundles ImageJ with a lot of plugins (including the 3 mentioned
above).
Ciao,
Johannes
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------------------------------
Date: Fri, 18 Nov 2011 13:24:11 -0600
From: Martin Wessendorf <[log in to unmask]>
Subject: Re: Olympus Corp in financial trouble?
*****
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http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
*****
Michael et al--
On 11/18/2011 11:28 AM, Cammer, Michael wrote:
> "Japanese officials say that at least $4.9 billion is unaccounted for in a financial scandal at Olympus<http://topics.nytimes.com/top/news/business/companies/olympus_corporation/index.html?inline=nyt-org> and are investigating whether much of that money went to companies with links to organized crime."
> http://www.nytimes.com/2011/11/18/business/global/japanese-police-investigate-olympus.html?_r=1&scp=3&sq=olympus&st=cse
I've seen these reports, too, over the past few weeks. However,
although it seems quite likely that their stockholders will be unhappy
with them, it's unclear (to me, at least!) that the company is in
financial trouble yet.
Disclaimer: I'm definitely not a certified financial advisor!
Martin Wessendorf
--
Martin Wessendorf, Ph.D. office: (612) 626-0145
Assoc Prof, Dept Neuroscience lab: (612) 624-2991
University of Minnesota Preferred FAX: (612) 624-8118
6-145 Jackson Hall, 321 Church St. SE Dept Fax: (612) 626-5009
Minneapolis, MN 55455 e-mail: [log in to unmask]
------------------------------
Date: Fri, 18 Nov 2011 15:26:43 -0600
From: Martin Wessendorf <[log in to unmask]>
Subject: (lots) more on Olympus
*****
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*****
For anyone who's interested:
http://newsfeedresearcher.com/data/articles_b47/olympus-woodford-company.html
Martin
--
Martin Wessendorf, Ph.D. office: (612) 626-0145
Assoc Prof, Dept Neuroscience lab: (612) 624-2991
University of Minnesota Preferred FAX: (612) 624-8118
6-145 Jackson Hall, 321 Church St. SE Dept Fax: (612) 626-5009
Minneapolis, MN 55455 e-mail: [log in to unmask]
------------------------------
Date: Fri, 18 Nov 2011 14:34:04 -0800
From: Iain Johnson <[log in to unmask]>
Subject: Re: Cell viability assay
*****
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*****
Calcein AM (intracellular esterase substrate). Incubate cells for 10
minutes in 1 microM calcein AM in serum-free medium at 37C. Wash off
staining medium and replace with complete culture medium. Image in
FITC/GFP channel.
You can also add a second dimension to the assay (at the expense of giving
up more spectral real estate).
SYTOX Orange (Molecular Probes; membrane permeability probe). Add to
complete culture medium at 5 microM. Do not wash. Image in TRITC/Cy3
channel.
Iain
Iain Johnson Consulting
Eugene, OR
(541) 285-8296
On Thu, Nov 17, 2011 at 7:23 AM, B. Prabhakar Pandian <[log in to unmask]> wrote:
> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/**wa?A0=confocalmicroscopy<http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy>
> *****
>
> Hello,
> I am emailing to see if anybody has a suggestion for a cell
> viability/toxicity assay that works similar to LDH/Caspase but relies on
> internal enzymes rather than in the supernatant. In addition, it should be
> compatible with fluorescence microscope.
>
> Thanks,
>
> -Prabhakar
>
------------------------------
Date: Fri, 18 Nov 2011 19:50:53 -0500
From: Christian Wilms <[log in to unmask]>
Subject: Re: Fast two-photon imaging: Constant fraction discriminator and variable delay for Laser pulse-synch
*****
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*****
Regarding the delay: if you do not need it to flexible in timing,
simply getting a coaxial cable of the correct length (20 cm
corresponds roughly to 1 ns) is one option. A bit more flexible and
still affordable are variable delay lines such as the Ortec 425 <http://www.ortec-online.com/download/425A.pdf
Best, Christian
------------------------------
Date: Fri, 18 Nov 2011 23:08:24 -0500
From: "Alison J. North" <[log in to unmask]>
Subject: Re: Olympus Corp in financial trouble? We certainly hope not!
*****
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*****
Hey Michael,
There is no question that Olympus has been going through hell this year - s=
tarting with the tsunami, and who could have guessed that things could have=
got even worse because of a few crooks (worst case) or idiots (best case) =
at the top.... Given that we own several high-end Olympus microscopes, we=
obviously have a vested interest in the company staying afloat and are und=
erstandably all concerned. More than that though, I have worked with the s=
ame Olympus team for over 11 years now and they have not only proved to be =
great colleagues but also good friends - we have caught wild mice on our ha=
nds and knees in my lab together, I have played despicable April Fool's jok=
es on them (well truthfully, only once - they have refused to visit my lab =
on April 1st ever since), and they have proved themselves such wizards at s=
olving the most mysterious and intractable microscope problems that we nick=
named the two local guys Dumbledore and Gandalf. At this point I think any=
discussion of what will happen is just pointless speculation, and one whic=
h they cannot possibly enter into, so the best we can do is to trust this w=
ill all blow over eventually and offer the microscope guys our support, say=
ing: we're rooting for you, and here's to 2012 being a far better year for =
you than 2011!
Best,
Alison
P.S. Sorry if I'm gushing to the confocal listserver - blame the glass of =
wine in my hand and the fact that I just came back from the movies :-).=20
=20
________________________________________
From: Confocal Microscopy List [[log in to unmask]] On Behalf=
Of Cammer, Michael [[log in to unmask]]
Sent: Friday, November 18, 2011 12:28 PM
To: [log in to unmask]
Subject: Olympus Corp in financial trouble?
*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=3Dconfocalmicroscopy
*****
"Japanese officials say that at least $4.9 billion is unaccounted for in a =
financial scandal at Olympus<http://topics.nytimes.com/top/news/business/co=
mpanies/olympus_corporation/index.html?inline=3Dnyt-org> and are investigat=
ing whether much of that money went to companies with links to organized cr=
ime."
http://www.nytimes.com/2011/11/18/business/global/japanese-police-investiga=
te-olympus.html?_r=3D1&scp=3D3&sq=3Dolympus&st=3Dcse
________________________________________________________
Michael Cammer, Assistant Research Scientist
Skirball Institute of Biomolecular Medicine
Lab: (212) 263-3208 Cell: (914) 309-3270
</PRE>
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d this email in error please notify the sender by return email and delete t=
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>
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------------------------------
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