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labeling cell nuclei

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Badri Roysam

labeling cell nuclei

Dear Colleagues, I am looking for a comparative review of methods for fluorescently labeling cell nuclei
for confocal imaging. Has anyone written such a review?

Thanks for any pointers,

Badri Roysam
Professor, Department of Electrical, Computer and Systems Engineering
Associate Director, NSF Center for Subsurface Sensing & Imaging Systems (CenSSIS ERC)
Co-Director, Rensselaer Center for Open Source Software
Rensselaer Polytechnic Institute
110 8th Street, Troy, New York 12180-3590, USA.
Office(JEC 7010): 518-276-8067, Assistant: 518-276-8525, Lab(JEC 6308): 518-276-8207, Fax: 518-276-8715
Email: [hidden email], Web: http://www.ecse.rpi.edu/~roysam

mahogny

Re: labeling cell nuclei

I'd like to know too. we found in our recent study that c.elegans his::RFP mutants accumulate random phenotypes, and other people have unofficially confirmed it with independently generated lines. someone really need to take a closer look at this issue.

<a href="http://dx.doi.org/10.1016/j.ydbio.2009.06.014" target="doilink" onclick="var doiWin; doiWin=window.open('http://dx.doi.org/10.1016/j.ydbio.2009.06.014','doilink','scrollbars=yes,resizable=yes,directories=yes,toolbar=yes,menubar=yes,status=yes'); doiWin.focus()">doi:10.1016/j.ydbio.2009.06.014

/Johan

On Fri, Jul 10, 2009 at 6:05 PM, Badri Roysam <[hidden email]> wrote:

Dear Colleagues, I am looking for a comparative review of methods for fluorescently labeling cell nuclei
for confocal imaging. Has anyone written such a review?

Thanks for any pointers,

Badri Roysam
Professor, Department of Electrical, Computer and Systems Engineering
Associate Director, NSF Center for Subsurface Sensing & Imaging Systems (CenSSIS ERC)
Co-Director, Rensselaer Center for Open Source Software
Rensselaer Polytechnic Institute
110 8th Street, Troy, New York 12180-3590, USA.
Office(JEC 7010): 518-276-8067, Assistant: 518-276-8525, Lab(JEC 6308): 518-276-8207, Fax: 518-276-8715
Email: [hidden email], Web: http://www.ecse.rpi.edu/~roysam

--
-----------------------------------------------------------
Johan Henriksson
PhD student, Karolinska Institutet
http://mahogny.areta.org http://www.endrov.net

Farid Jalali

Re: labeling cell nuclei

In reply to this post by Badri Roysam

Hello Badri,

A couple of these are quite all-encompassing. The article by Wojcik and Dobrucki will be of particular interest if you are looking at chromatin dynamics and processes that impact chromatin structure.

Hope it helps.
Cheers
Farid

1. Stuart KR and ES Cole. Nuclear and cytoskeletal fluorescence microscopy techniques. Methods in Cell Biology (2001). 62:291-311

2. Ploeger LS, Dullens HF, Huisman A, van Diest PJ. Fluorescent stains for quantification of DNA by confocal laser scanning microscopy in 3-D. Biotechnic Histochemistry (2008). 83(2):63-9.

3. Suzuki T, Fujikura K, Higashiyama T, Takata K. DNA Staining for fluorescence and laser confocal microscopy. Journal of Histochemistry and Cytochemistry (1997). 45(1):49-53.

4. Wojcik K and JW Dobrucki. Interaction of a DNA intercalator DRAQ5, and a minor groove binder SYTO17, with chromatin in live cells--influence on chromatin organization and histone-DNA interactions. Cytometry A (2008). 73(6):555-562.

On Fri, Jul 10, 2009 at 9:05 AM, Badri Roysam <[hidden email]> wrote:

Dear Colleagues, I am looking for a comparative review of methods for fluorescently labeling cell nuclei
for confocal imaging. Has anyone written such a review?

Thanks for any pointers,

Badri Roysam
Professor, Department of Electrical, Computer and Systems Engineering
Associate Director, NSF Center for Subsurface Sensing & Imaging Systems (CenSSIS ERC)
Co-Director, Rensselaer Center for Open Source Software
Rensselaer Polytechnic Institute
110 8th Street, Troy, New York 12180-3590, USA.
Office(JEC 7010): 518-276-8067, Assistant: 518-276-8525, Lab(JEC 6308): 518-276-8207, Fax: 518-276-8715
Email: [hidden email], Web: http://www.ecse.rpi.edu/~roysam

Keith Morris

Re: labeling cell nuclei

Hi Badri,

I assume we are talking molecular cytogenetics. Often confocals aren’t the first choice at imaging FISH [fluorescent in-situ hybridization] metaphase chromosomes & interphase nuclei, although we do use ours for 3D FISH, i.e. looking at chromatin organization in the nuclei in, naturally, 3D [see Farids post]. Otherwise most molecular cytogenetics systems involve 100x [even 150x] high NA oil objectives and a standard wide-field upright fluorescence microscope [with multiple narrow band fluorescence filter sets]. These microscopes are controlled via specialized cytogenetics software* for karyotyping, CGH, mFISH etc... The nuclei/chromosomes are typically stripped of the surrounding cell cytoplasm and hence quite flat [and the structures are very small]. Imaging is via specific fluorescence chromosome/centromere/telomere/arm paints and probes targeted to small sequences of DNA. My line manager [or ‘boss’ as we used to say] is in fact in the process of editing a large tome on all things FISH related, and this should be published early next year.

Regards

Keith

Links to molecular cytogenetics websites
http://www.well.ox.ac.uk/cytogenetics/websites.shtml

Reading list
http://www.well.ox.ac.uk/cytogenetics/reading.shtml

*e.g. Applied Imaging [Genetix]
http://www.genetix.com/en/systems/cytovision/introduction/index.html
Metasystems
http://www.metasystems.de/indexold.htm

Spectral-imaging

http://www.spectral-imaging.com/spectral.asp

---------------------------------------------------------------------------
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/

From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Farid Jalali
Sent: 10 July 2009 22:34
To: [hidden email]
Subject: Re: labeling cell nuclei

On Fri, Jul 10, 2009 at 9:05 AM, Badri Roysam <[hidden email]> wrote:

Badri Roysam

Re: labeling cell nuclei

In reply to this post by Badri Roysam

Monica K. Chawla, Gang Lin, Kathy Olson, Almira Vazdarjanova, S. N. Burke, B. L. McNaughton, P. F. Worley, John F. Guzowski, Badrinath Roysam and C. A. Barnes, “3D-catFISH: A System for Automated Quantitative Three-Dimensional Compartmental Analysis of Temporal Gene Transcription Activity Imaged by Fluorescence in situ Hybridization,” Journal of Neuroscience Methods, 139, 13–24, 2004.

John F. Guzowski, Jerilyn A. Timlin, Badri Roysam, Bruce L. Mc Naughton, Paul F. Worley and Carol A. Barnes, “Mapping Behaviorally Relevant Neural Circuits With Immediate Early gene Expression,” Current Opinion in Neurobiology, Vol 15, Issue 5, 2005.

Vicki L Sutherland, Jerilyn A Timlin, Linda T Nieman, John F Guzowski, Monica K Chawla, Paul F Worley, Badri Roysam, Bruce L McNaughton, Michael B Sinclair, Carol A Barnes, “Advanced Imaging of Multiple mRNAs in Brain Tissue using a Custom Hyperspectral Imager and Multivariate Curve Resolution,” Journal of Neuroscience Methods, Feb 15;160(1):144-8, 2007.

Thanks,

Badri Roysam
Professor, Department of Electrical, Computer and Systems Engineering
Associate Director, NSF Center for Subsurface Sensing & Imaging Systems (CenSSIS ERC)
Co-Director, Rensselaer Center for Open Source Software
Rensselaer Polytechnic Institute
110 8th Street, Troy, New York 12180-3590, USA.
Office(JEC 7010): 518-276-8067, Assistant: 518-276-8525, Lab(JEC 6308): 518-276-8207, Fax: 518-276-8715
Email: [hidden email], Web: http://www.ecse.rpi.edu/~roysam

----- Original Message -----
From: Keith Morris [mailto:[hidden email]]
To: [hidden email]
Subject: Re: labeling cell nuclei

> Hi Badri,
>
>
>
> I assume we are talking molecular cytogenetics. Often confocals aren't the
> first choice at imaging FISH [fluorescent in-situ hybridization] metaphase
> chromosomes & interphase nuclei, although we do use ours for 3D FISH, i.e.
> looking at chromatin organization in the nuclei in, naturally, 3D [see
> Farids post]. Otherwise most molecular cytogenetics systems involve 100x
> [even 150x] high NA oil objectives and a standard wide-field upright
> fluorescence microscope [with multiple narrow band fluorescence filter
> sets]. These microscopes are controlled via specialized cytogenetics
> software* for karyotyping, CGH, mFISH etc... The nuclei/chromosomes are
> typically stripped of the surrounding cell cytoplasm and hence quite flat
> [and the structures are very small]. Imaging is via specific fluorescence
> chromosome/centromere/telomere/arm paints and probes targeted to small
> sequences of DNA. My line manager [or 'boss' as we used to say] is in fact
> in the process of editing a large tome on all things FISH related, and this
> should be published early next year.
>
>
>
> Regards
>
>
>
> Keith
>
>
>
> Links to molecular cytogenetics websites
> http://www.well.ox.ac.uk/cytogenetics/websites.shtml
>
> Reading list
> http://www.well.ox.ac.uk/cytogenetics/reading.shtml
>
>
>
>
> *e.g. Applied Imaging [Genetix]
> http://www.genetix.com/en/systems/cytovision/introduction/index.html
> Metasystems
> http://www.metasystems.de/indexold.htm
>
> Spectral-imaging
>
> http://www.spectral-imaging.com/spectral.asp
>
>
>
>
>
>
>
> ---------------------------------------------------------------------------
> 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: <mailto:[hidden email]> [hidden email]
> Web-pages: <http://www.well.ox.ac.uk/cytogenetics/>
> http://www.well.ox.ac.uk/cytogenetics/
>
> From: Confocal Microscopy List [mailto:[hidden email]] On
> Behalf Of Farid Jalali
> Sent: 10 July 2009 22:34
> To: [hidden email]
> Subject: Re: labeling cell nuclei
>
>
>
> Hello Badri,
>
> A couple of these are quite all-encompassing. The article by Wojcik and
> Dobrucki will be of particular interest if you are looking at chromatin
> dynamics and processes that impact chromatin structure.
>
> Hope it helps.
> Cheers
> Farid
>
> 1. Stuart KR and ES Cole. Nuclear and cytoskeletal fluorescence microscopy
> techniques. Methods in Cell Biology (2001). 62:291-311
>
> 2. Ploeger LS, Dullens HF, Huisman A, van Diest PJ. Fluorescent stains for
> quantification of DNA by confocal laser scanning microscopy in 3-D.
> Biotechnic Histochemistry (2008). 83(2):63-9.
>
> 3. Suzuki T, Fujikura K, Higashiyama T, Takata K. DNA Staining for
> fluorescence and laser confocal microscopy. Journal of Histochemistry and
> Cytochemistry (1997). 45(1):49-53.
>
> 4. Wojcik K and JW Dobrucki. Interaction of a DNA intercalator DRAQ5, and a
> minor groove binder SYTO17, with chromatin in live cells--influence on
> chromatin organization and histone-DNA interactions. Cytometry A (2008).
> 73(6):555-562.
>
>
>
>
> On Fri, Jul 10, 2009 at 9:05 AM, Badri Roysam <[hidden email]> wrote:
>
> Dear Colleagues, I am looking for a comparative review of methods for
> fluorescently labeling cell nuclei
> for confocal imaging. Has anyone written such a review?
>
> Thanks for any pointers,
>
>
> Badri Roysam
> Professor, Department of Electrical, Computer and Systems Engineering
> Associate Director, NSF Center for Subsurface Sensing & Imaging Systems
> (CenSSIS ERC)
> Co-Director, Rensselaer Center for Open Source Software
> Rensselaer Polytechnic Institute
> 110 8th Street, Troy, New York 12180-3590, USA.
> Office(JEC 7010): 518-276-8067, Assistant: 518-276-8525, Lab(JEC 6308):
> 518-276-8207, Fax: 518-276-8715
> Email: [hidden email], Web: http://www.ecse.rpi.edu/~roysam
> <http://www.ecse.rpi.edu/%7Eroysam>
>
>
>
>

Keith Morris

Re: labeling cell nuclei

Hi Badri,

Thanks for the article links. As you can see from the published images in these papers, the confocal scans presented have a tendency to be a little fuzzy compared to a dedicated molecular cytogenetics microscope system, not that the image 'imformation' is compromised.

Keith

---------------------------------------------------------------------------
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 Badri Roysam
Sent: 16 July 2009 11:22
To: [hidden email]
Subject: Re: labeling cell nuclei

Monica K. Chawla, Gang Lin, Kathy Olson, Almira Vazdarjanova, S. N. Burke, B. L. McNaughton, P. F. Worley, John F. Guzowski, Badrinath Roysam and C. A. Barnes, “3D-catFISH: A System for Automated Quantitative Three-Dimensional Compartmental Analysis of Temporal Gene Transcription Activity Imaged by Fluorescence in situ Hybridization,” Journal of Neuroscience Methods, 139, 13–24, 2004.

John F. Guzowski, Jerilyn A. Timlin, Badri Roysam, Bruce L. Mc Naughton, Paul F. Worley and Carol A. Barnes, “Mapping Behaviorally Relevant Neural Circuits With Immediate Early gene Expression,” Current Opinion in Neurobiology, Vol 15, Issue 5, 2005.

Vicki L Sutherland, Jerilyn A Timlin, Linda T Nieman, John F Guzowski, Monica K Chawla, Paul F Worley, Badri Roysam, Bruce L McNaughton, Michael B Sinclair, Carol A Barnes, “Advanced Imaging of Multiple mRNAs in Brain Tissue using a Custom Hyperspectral Imager and Multivariate Curve Resolution,” Journal of Neuroscience Methods, Feb 15;160(1):144-8, 2007.

Thanks,

Badri Roysam
Professor, Department of Electrical, Computer and Systems Engineering
Associate Director, NSF Center for Subsurface Sensing & Imaging Systems (CenSSIS ERC)
Co-Director, Rensselaer Center for Open Source Software
Rensselaer Polytechnic Institute
110 8th Street, Troy, New York 12180-3590, USA.
Office(JEC 7010): 518-276-8067, Assistant: 518-276-8525, Lab(JEC 6308): 518-276-8207, Fax: 518-276-8715
Email: [hidden email], Web: http://www.ecse.rpi.edu/~roysam

----- Original Message -----
From: Keith Morris [mailto:[hidden email]]
To: [hidden email]
Subject: Re: labeling cell nuclei