http://confocal-microscopy-list.275.s1.nabble.com/Decon-wall-tp7580700p7580706.html
As for the software, it's SoftWorx. But I have been playing around with
ImageJ to troubleshoot because it's faster. The cutting off problem only
occurs when I deconvolve with apply correction in SoftWorx. I don't see it
Tom: Thanks! I'll check those out immediately. You're right that the signal
> *****
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>
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> *****
>
> Heather,
>
> to get some help from the list members it would be helpful if you'd let us
> know in which corner of the world you are located. Your G-mail account
> doesn't give a clue and you don't have a signature. I get my
> non-fluorescent coverslips from
http://www.hecht-assistent.de/ (Order
> no.:1014) but I am not sure if you would consider ordering in Germany.
>
> Also, you might want to let people know which decon software you are using.
>
> Steffen
>
> On 16.07.2013 06:23, Heather Bowden wrote:
>
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>>
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>>
>>
>> Dear Confocal Microscopy List,
>>
>> I have hit another wall with a very old deconvolution microscope (late
>> 90's).
>>
>> I have images from fixed slides that have DAPI and a green dye. I grow
>> the
>> cells on round coverslips that are pre-coated with Poly-D-Lysine or
>> Poly-L-Lysine from the supplier (BD).
>>
>> 1) I'm having issues with green autofluorescence. As a result, my
>> deconvolved images don't look very good. They are very grainy and almost
>> look homogeneous. I'm confident it's the coverslip that's causing it but I
>> can't buy them without the poly-lysine and I can't grow the cells without
>> poly-lysine either. I tried a round Fisher coverslip with no coating, but
>> that had autofluorescence too. I have been using extra long rectangular
>> coverslips for mounting because those don't have autofluorescence. Any
>> suggestions would be incredibly appreciated.
>>
>> 2) When I deconvolve a slide I made, it cuts off the right side of the
>> image and places it on the left. There's usually a big line where it made
>> the cut. Why would this occur? When I use a prepared fixed slide from
>> Invitrogen (Fluocell #2), it cuts off the left side and puts it on the
>> right! This only happens when I deconvolve wavelengths separately. I do
>> not
>> touch the "pass wave" function (what does that do?).
>>
>> 3) What glass slides do you use? Cat #'s would be greatly appreciated as
>> the ones Invitrogen recommended aren't available anymore and Fisher does
>> not know (and their slides autofluoresce).
>>
>> Thank you so much for any help!
>>
>> Heather
>>
>>
>
> --
> ------------------------------**------------------------------
> Steffen Dietzel, PD Dr. rer. nat
> Ludwig-Maximilians-Universität München
> Walter-Brendel-Zentrum für experimentelle Medizin (WBex)
> Head of light microscopy
>
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