Mowiol and 3D imaging

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ChrisWilms ChrisWilms
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Mowiol and 3D imaging

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Dear List,

I've read that Mowiol is not considered well suited for 3D imaging:
<http://www.biocenter.helsinki.fi/bi/lmu/images/Mountants.pdf>

Unfortunately, the authors give no specifics on why that should be the  
case and I also can't find anything on the web. Does anyone have  
details on this?

Thanks,

Chris
Mario Faretta Mario Faretta
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Re: Mowiol and 3D imaging

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Dear Christian,
the main problem with Mowiol and in general with all the mounting media that
become solid over time, is that there is a compression flattening cells. This
effect tend to be obviously worst as time goes on and it is particularly
dramatic in cell monolayers (I tried just for fun to measure nucleus thickness
after 1 week and more than two weeks: staining was perfectly maintained but
cell thickness reduced of more than 50% in 1 week and goes down up to few
microns at the end (at least at the concentration we use)). The point is that
if you need to use confocal generally you do it for a 3D analysis but if you
prepare the sample this way you make it 2D. Depending on the goal of the
experiment Mowiol can be deleterious almost immediately: I remember an old
pubblication in Cytometry from Tom Jovin (hope I am correct) showing apical
and basal membrane fusion at the cell periphery due to mounting media induced
compression...
hope it helps
Mario

 Christian Wilms ([hidden email]) wrote:
 >
 > *****
 > To join, leave or search the confocal microscopy listserv, go to:
 > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
 > *****
 >
 > Dear List,
 >
 > I've read that Mowiol is not considered well suited for 3D imaging:
 >
 >
 > Unfortunately, the authors give no specifics on why that should be the
 > case and I also can't find anything on the web. Does anyone have
 > details on this?
 >
 > Thanks,
 >
 > Chris
 >
Patrick Van Oostveldt Patrick Van Oostveldt
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Re: Mowiol and 3D imaging

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Dear,

You also have to take into account that the refractive index of the  
mounting medium can be changed due to solidification.
An increase in RI will induce measuring errors cfr: Visser and  
Braekenhoff some years a go in J.Microscopy and the measured thickness  
will be reduced.



Greetings

Patrick Van Oostveldt

Quoting "Mario Faretta" <[hidden email]>:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> *****
>
> Dear Christian,
> the main problem with Mowiol and in general with all the mounting media that
> become solid over time, is that there is a compression flattening cells. This
> effect tend to be obviously worst as time goes on and it is particularly
> dramatic in cell monolayers (I tried just for fun to measure nucleus  
> thickness
> after 1 week and more than two weeks: staining was perfectly maintained but
> cell thickness reduced of more than 50% in 1 week and goes down up to few
> microns at the end (at least at the concentration we use)). The point is that
> if you need to use confocal generally you do it for a 3D analysis but if you
> prepare the sample this way you make it 2D. Depending on the goal of the
> experiment Mowiol can be deleterious almost immediately: I remember an old
> pubblication in Cytometry from Tom Jovin (hope I am correct) showing apical
> and basal membrane fusion at the cell periphery due to mounting media induced
> compression...
> hope it helps
> Mario
>
>  Christian Wilms ([hidden email]) wrote:
>  >
>  > *****
>  > To join, leave or search the confocal microscopy listserv, go to:
>  > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>  > *****
>  >
>  > Dear List,
>  >
>  > I've read that Mowiol is not considered well suited for 3D imaging:
>  >
>  >
>  > Unfortunately, the authors give no specifics on why that should be the
>  > case and I also can't find anything on the web. Does anyone have
>  > details on this?
>  >
>  > Thanks,
>  >
>  > Chris
>  >
>



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ChrisWilms ChrisWilms
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Re: Mowiol and 3D imaging

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*****
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Dear all,

thanks for your responses on this. Sounds as if Mowiol has the same  
problems as most other low RI and/or curing mountants.

As far as I gathered, the RI of Mowiol is too low for good axial  
measurements even when solidified. I'm still trying to chase down a  
mounting medium with a RI similar to immersion oil that doesn't quench  
fluorescence excessively. I've tried TDE and actual immersion oil --  
both gave pretty dim images in contrast to other mounting media  
(Vectashield, Prolong). I'm now trying several of Citifluor's, hoping  
to find a good mix.

Again, thanks for the responses!

Best, Christian