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Hi Christophe,
Prolong Gold: "20x of almost nothing is almost nothing". I charge $45/hr
for access to the Leica SP5 inverted confocal microscope I manage (and
we'll probably reach ~1,000 hours, enough to cover its service contract
charge ... 9 laser lines, motorized stage, galvo-Z). If your microscope
has similar value, and you spend one hour per slide, the cost of Prolong
Gold is miniscule compared to the confocal rate + cost of your time to
make the slide and do the imaging. I strongly recommend Prolong Gold
without DAPI, low concentration of DAPI after the last antibody wash
step. Lots of antibody wash steps (7 or more), paying attention not to
let any specimen dry (that is, remove/add liquid to one coverglass).
***
You might try combining the listed ingredients from the item below:
http://www.aname.es/microscopia/ems/chemicals/adhesive.htmlFluoromount-G Slide Mounting Medium
A special mounting media for immunofluorescent staining
procedures. It is a water soluble, non-fluorescing medium which is well
suited for use when the staining procedure has an aqueous final step. It
contains 10% polyvinyl alcohol in phosphate buffered
glycerol with 0.1% sodium azide as a preservative. One bottle is
sufficient to mount up to 500 slides.
0-5ºC 17984-25
If you feel adventurous, you might try combining PVA with
2,2'-thiodiethanol ... look up Stan Vitha's method for gradual
infiltration of TDE (transition from aqueous to TDE). Note; I have no
idea if PVA will solidify in mostly TDE (will not be surprised if it
does work).
If you need more ideas, see
http://www.microscopy-uk.org.uk/mag/artjun04/wdmtmedia.html (not
necessarily intended for fluorescence).
George
On 10/24/2011 9:13 AM, Christophe Leterrier wrote:
> *****
> To join, leave or search the confocal microscopy listserv, go to:
>
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy> *****
>
> Hi,
>
> I was a happy user of Fluorsave for mounting immunlabeled cells cultured on
> coverlips. Recently though, our Fluorsave has stopped hardening after
> overnight curing at RT (it has happened with our regular Fluorsave and with
> another bottle from a different lot we ordered after this problem
> appeared).
>
> First, is there any user of Fluorsave that has had this problem and managed
> to find an explanation/solution to it?
> Second, can you recommend a good alternative for a mounting cultured cells
> labelled with Alexa/DyLight fluorophores ?
>
> I'd prefer something that does not require sealing. I had a look at Tony
> Collin's "Mountants" document (
>
http://www.lajollaneuroscience.org/sr/homepage/cell/Mountants.pdf) and tried
> ProLong Gold, which is very nice but too expensive for routine work
> (something like 20 times the price of Fluorsave). I got some FluoroGel (
>
http://www.emsdiasum.com/microscopy/technical/datasheet/17985.aspx) and I'm
> currently evaluating it.
>
> Any other advice or mountant proposition ?
>
> Thanks,
>
>
> --
> Christophe Leterrier
> Postdoc
> INSERM UMR641 Neurobiologie des Canaux Ioniques
> IFR Jean Roche, Université de la Méditerranée
> Marseille, France
>
[hidden email]
>
>
--
George McNamara, PhD
Analytical Imaging Core Facility
University of Miami
Locked
