Dyes in the far red spectrum

[Matt Wayland]

*****
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,

We are looking to do 5-channel immuno-fluorescence imaging on our Leica
SP5 confocal equipped with the following excitation wavelengths: 405,
488, 561, 633nm. And we were hoping to use the following fluorophores:
CF405, Alexa488, CF568, CF633.

Are there any good far red fluorophores that are excited at 633nm and
which might be sufficiently distinct from CF633? Or are there other
combinations of fluorophores that would work well for 5-channel imaging?

Many thanks in advance,

Matt

--
Matt Wayland
Imaging Facility Manager
Department of Zoology
University of Cambridge
Downing Street
Cambridge
CB2 3EJ

Tel.: (++44) (0)1223 769282
Fax.: (++44) (0)1223 336676

[George McNamara]

*****
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.
*****

Hi Matt,

At

http://www.geomcnamara.com/fluorophore-table

the columns are supposed to be sortable.

Table is a bit out of date since Janelia Fluor 646 is not shown. Contact
Luke Lavis, HHMI JRC for JF fluorophores:

https://www.janelia.org/open-science/janelia-fluor-dyes

The following dyes are available for Non-Profit research for free:

  * Janelia Fluor™ 549
  * Janelia Fluor™ 646
  * Photoactivatible Janelia Fluor™ 549
  * Photoactivatible Janelia Fluor™ 646

These dyes and patent rights are available for Commercial License for
the full color spectrum as well.

Janelia Fluor™ 549
<https://www.tocris.com/dispprod.php?ItemId=524906#.WLXV0hIrKg8>andJanelia Fluor™ 646
<https://www.tocris.com/dispprod.php?ItemId=524907#.WLXV5BIrKg8>are
available fromTocris Bioscience <https://www.tocris.com/> for purchase.


//

Consider going Brilliant (BD Biosciences), i.e. 405 nm excitation ->
6plex (including BV711 and BV785, see also BUVs, BBs) or Super Bright
(Thermo Fisher / eBioscience / etc), see

https://www.linkedin.com/pulse/bd-biosciences-listed-tandems-horizon-brilliant-violets-mcnamara

Semrock, Chroma, etc, have spectra graphing of the Brilliant's.
ThermoFisher has Super Brights (but not Brilliant's).

https://www.thermofisher.com/us/en/home/life-science/cell-analysis/labeling-chemistry/fluorescence-spectraviewer.html

Most of the graphing sites have download capability. Older fluorophores
spectra are in the PubSpectra XLSX file at

https://works.bepress.com/gmcnamara/9/

enjoy,

George



On 3/1/2017 11:26 AM, Matt Wayland 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,
>
> We are looking to do 5-channel immuno-fluorescence imaging on our
> Leica SP5 confocal equipped with the following excitation wavelengths:
> 405, 488, 561, 633nm. And we were hoping to use the following
> fluorophores: CF405, Alexa488, CF568, CF633.
>
> Are there any good far red fluorophores that are excited at 633nm and
> which might be sufficiently distinct from CF633? Or are there other
> combinations of fluorophores that would work well for 5-channel imaging?
>
> Many thanks in advance,
>
> Matt
>

--


George McNamara, PhD
Houston, TX 77054
[hidden email]
https://www.linkedin.com/in/georgemcnamara
https://works.bepress.com/gmcnamara/75/
http://www.ncbi.nlm.nih.gov/myncbi/browse/collection/44962650

[Romain Laine]

*****
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.
*****

Hi Matt,
I know a handful of people have used Alexa Fluor 700 and 647 together
excited with a 640 nm laser.
See here a paper from the Sauer group.

http://www.nature.com/nmeth/journal/v9/n11/abs/nmeth.2224.html

You need the right filters and will probably need to do some unmixing
though.

Cheers,

Romain


--
Dr. Romain Laine, PhD in Biophotonics
Laser Analytics Group
Department of Chemical Engineering and Biotechnology
University of Cambridge
West Cambridge Site
Philippa Fawcett Drive
Cambridge
CB3 0AS


On Wed, Mar 1, 2017 at 5:26 PM, Matt Wayland <[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 All,
>
> We are looking to do 5-channel immuno-fluorescence imaging on our Leica
> SP5 confocal equipped with the following excitation wavelengths: 405, 488,
> 561, 633nm. And we were hoping to use the following fluorophores: CF405,
> Alexa488, CF568, CF633.
>
> Are there any good far red fluorophores that are excited at 633nm and
> which might be sufficiently distinct from CF633? Or are there other
> combinations of fluorophores that would work well for 5-channel imaging?
>
> Many thanks in advance,
>
> Matt
>
> --
> Matt Wayland
> Imaging Facility Manager
> Department of Zoology
> University of Cambridge
> Downing Street
> Cambridge
> CB2 3EJ
>
> Tel.: (++44) (0)1223 769282
> Fax.: (++44) (0)1223 336676
>