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green nuclear stain

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Sylvie Le Guyader-2

green nuclear stain

Hi there!

Does anyone know of a green nuclear stain that resists fixation? One of our users wants to do some colocalization of cy3 and cy5 with the nucleus and our confocal has unfortunately no 405 laser. We are trying to avoid NLS-GFP but use something that would label all cells right before or after fixation.

Thanks to all!

Med vänlig hälsning / Best regards

Sylvie

@@@@@@@@@@@@@@@@@@@@@@@@

Sylvie Le Guyader

Dept of Biosciences and Nutrition

Karolinska Institutet

Novum

14157 Huddinge

Sweden

+46 (0)8 608 9240

Cameron Nowell

Re: green nuclear stain

Hi Sylvie,

I have had great success in the past using OliGreen from Invitrogen (no commercial affiliation), though it is not supplied specifically as a DNA stain.

Cheers

Cam

Cameron J. Nowell
Microscpy Manager
Central Resource for Advanced Microscopy
Ludwig Insttue for Cancer Research
PO Box 2008
Royal Melbourne Hospital
Victoria, 3050
AUSTRALIA

Office: +61 3 9341 3155
Mobile: +61422882700
Fax: +61 3 9341 3104

http://www.ludwig.edu.au/branch/research/platform/microscopy.htm

________________________________

From: Confocal Microscopy List on behalf of Sylvie Le Guyader
Sent: Wed 30/09/2009 7:20 PM
To: [hidden email]
Subject: green nuclear stain

Hi there!

Does anyone know of a green nuclear stain that resists fixation? One of our users wants to do some colocalization of cy3 and cy5 with the nucleus and our confocal has unfortunately no 405 laser. We are trying to avoid NLS-GFP but use something that would label all cells right before or after fixation.

Thanks to all!

Med vänlig hälsning / Best regards

Sylvie

@@@@@@@@@@@@@@@@@@@@@@@@

Sylvie Le Guyader

Dept of Biosciences and Nutrition

Karolinska Institutet

Novum

14157 Huddinge

Sweden

+46 (0)8 608 9240

anh2006

Re: green nuclear stain

In reply to this post by Sylvie Le Guyader-2

Yopro from Invitrogen is a green fluorescent nuclear counterstain.

From: Sylvie Le Guyader
Date: Wed, 30 Sep 2009 11:20:08 +0200
To: <[hidden email]>
Subject: green nuclear stain

Hi there!

Thanks to all!

Med vänlig hälsning / Best regards

Sylvie

@@@@@@@@@@@@@@@@@@@@@@@@

Sylvie Le Guyader

Dept of Biosciences and Nutrition

Karolinska Institutet

Novum

14157 Huddinge

Sweden

+46 (0)8 608 9240

Michael Weber-4

Re: green nuclear stain

In reply to this post by Sylvie Le Guyader-2

Tjänare Sylvie,

you might try Sytox Green or TO-PRO 1.

Michael

> Hi there!
>
>
>
> Does anyone know of a green nuclear stain that resists fixation? One of
> our
> users wants to do some colocalization of cy3 and cy5 with the nucleus and
> our confocal has unfortunately no 405 laser. We are trying to avoid
> NLS-GFP
> but use something that would label all cells right before or after
> fixation.
>
>
>
> Thanks to all!
>
>
>
> Med vänlig hälsning / Best regards
>
>
>
> Sylvie
>
>
>
> @@@@@@@@@@@@@@@@@@@@@@@@
>
> Sylvie Le Guyader
>
> Dept of Biosciences and Nutrition
>
> Karolinska Institutet
>
> Novum
>
> 14157 Huddinge
>
> Sweden
>
> +46 (0)8 608 9240

Michael Weber-4

Re: green nuclear stain

In reply to this post by Sylvie Le Guyader-2

Forgot to say... for colocalization with Cy3 I would prefer the Sytox
Green, unless you are using a 561 nm laser for Cy3. The TO-PRO 1 is still
excitable with 543 nm and will therefore bleed into Cy3 which for sure
messes up the analysis.

Michael

Judy Trogadis

Re: green nuclear stain

In reply to this post by Sylvie Le Guyader-2

Sytox Green (Molecular Probes) works well for fixed, not sure about live
cells. Normal dilution is 1:20,000
but titrate because it is very bright and depending on your setup and
other dyes may be seen in other channels unless it is at max dilution.

Judy

Judy Trogadis
Bio-Imaging Coordinator
St. Michael's Hospital, 7Queen
30 Bond St.
Toronto, ON M5B 1W8, Canada
ph: 416-864-6060 x6337
pager: 416-685-9219
fax: 416-864-5046
[hidden email]

>>> Sylvie Le Guyader <[hidden email]> 9/30/2009 5:20 AM >>>
Hi there!

Does anyone know of a green nuclear stain that resists fixation? One of
our
users wants to do some colocalization of cy3 and cy5 with the nucleus
and
our confocal has unfortunately no 405 laser. We are trying to avoid
NLS-GFP
but use something that would label all cells right before or after
fixation.

Thanks to all!

Med vänlig hälsning / Best regards

Sylvie

@@@@@@@@@@@@@@@@@@@@@@@@

Sylvie Le Guyader

Dept of Biosciences and Nutrition

Karolinska Institutet

Novum

14157 Huddinge

Sweden

+46 (0)8 608 9240

Sylvie Le Guyader-2

Re: green nuclear stain

Dear all

Thanks a lot for all the suggestions. We will try and hopefully get nice
results!

:)

Med vänlig hälsning / Best regards

Sylvie

@@@@@@@@@@@@@@@@@@@@@@@@
Sylvie Le Guyader
Dept of Biosciences and Nutrition
Karolinska Institutet
Novum
14157 Huddinge
Sweden
+46 (0)8 608 9240

> -----Original Message-----
> From: Confocal Microscopy List
> [mailto:[hidden email]] On Behalf Of Judy
> Trogadis
> Sent: 30 September 2009 15:26
> To: [hidden email]
> Subject: Re: green nuclear stain
>
> Sytox Green (Molecular Probes) works well for fixed, not sure about live
> cells. Normal dilution is 1:20,000
> but titrate because it is very bright and depending on your setup and
> other dyes may be seen in other channels unless it is at max dilution.
>
> Judy
>
> Judy Trogadis
> Bio-Imaging Coordinator
> St. Michael's Hospital, 7Queen
> 30 Bond St.
> Toronto, ON M5B 1W8, Canada
> ph: 416-864-6060 x6337
> pager: 416-685-9219
> fax: 416-864-5046
> [hidden email]
>
>
> >>> Sylvie Le Guyader <[hidden email]> 9/30/2009 5:20 AM >>>
> Hi there!
>
>
>
> Does anyone know of a green nuclear stain that resists fixation? One of
> our
> users wants to do some colocalization of cy3 and cy5 with the nucleus
> and
> our confocal has unfortunately no 405 laser. We are trying to avoid
> NLS-GFP
> but use something that would label all cells right before or after
> fixation.
>
>
>
> Thanks to all!
>
>
>
> Med vänlig hälsning / Best regards
>
>
>
> Sylvie
>
>
>
> @@@@@@@@@@@@@@@@@@@@@@@@
>
> Sylvie Le Guyader
>
> Dept of Biosciences and Nutrition
>
> Karolinska Institutet
>
> Novum
>
> 14157 Huddinge
>
> Sweden
>
> +46 (0)8 608 9240
>
>

Steffen Dietzel

Re: green nuclear stain

Sylvie,

you also might want to have a look at Chromomycin A3.
disadvantage: needs Mg-Ions or will not stain
(you surely can google some protocols)
advantages: DNA-specific (most others also stain
RNA); old and patent free and thus comparably cheap (if I remember right).

Steffen

At 15:40 30.09.2009, you wrote:

>Dear all
>
>Thanks a lot for all the suggestions. We will try and hopefully get nice
>results!
>
>:)
>
>Med vänlig hälsning / Best regards
>
>Sylvie
>
>@@@@@@@@@@@@@@@@@@@@@@@@
>Sylvie Le Guyader
>Dept of Biosciences and Nutrition
>Karolinska Institutet
>Novum
>14157 Huddinge
>Sweden
>+46 (0)8 608 9240
>
> > -----Original Message-----
> > From: Confocal Microscopy List
> > [mailto:[hidden email]] On Behalf Of Judy
> > Trogadis
> > Sent: 30 September 2009 15:26
> > To: [hidden email]
> > Subject: Re: green nuclear stain
> >
> > Sytox Green (Molecular Probes) works well for fixed, not sure about live
> > cells. Normal dilution is 1:20,000
> > but titrate because it is very bright and depending on your setup and
> > other dyes may be seen in other channels unless it is at max dilution.
> >
> > Judy
> >
> > Judy Trogadis
> > Bio-Imaging Coordinator
> > St. Michael's Hospital, 7Queen
> > 30 Bond St.
> > Toronto, ON M5B 1W8, Canada
> > ph: 416-864-6060 x6337
> > pager: 416-685-9219
> > fax: 416-864-5046
> > [hidden email]
> >
> >
> > >>> Sylvie Le Guyader <[hidden email]> 9/30/2009 5:20 AM >>>
> > Hi there!
> >
> >
> >
> > Does anyone know of a green nuclear stain that resists fixation? One of
> > our
> > users wants to do some colocalization of cy3 and cy5 with the nucleus
> > and
> > our confocal has unfortunately no 405 laser. We are trying to avoid
> > NLS-GFP
> > but use something that would label all cells right before or after
> > fixation.
> >
> >
> >
> > Thanks to all!
> >
> >
> >
> > Med vänlig hälsning / Best regards
> >
> >
> >
> > Sylvie
> >
> >
> >
> > @@@@@@@@@@@@@@@@@@@@@@@@
> >
> > Sylvie Le Guyader
> >
> > Dept of Biosciences and Nutrition
> >
> > Karolinska Institutet
> >
> > Novum
> >
> > 14157 Huddinge
> >
> > Sweden
> >
> > +46 (0)8 608 9240
> >
> >

Cameron Nowell

Re: green nuclear stain

HI all,

While i haven't used the dye Steffen recommends, i have used DNA dyes that light up RNA as well. Give a quick treatment of your samples with RNase prior to staining will get rid of the problem.

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 Steffen Dietzel
Sent: Thursday, 1 October 2009 11:05 PM
To: [hidden email]
Subject: Re: green nuclear stain

Sylvie,

you also might want to have a look at Chromomycin A3.
disadvantage: needs Mg-Ions or will not stain
(you surely can google some protocols)
advantages: DNA-specific (most others also stain
RNA); old and patent free and thus comparably cheap (if I remember right).

Steffen

At 15:40 30.09.2009, you wrote:

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