Fluorescence microscopy

MMu brilhasti at gmx.net
Mon Feb 7 06:35:55 EST 2005


"Wojtek" <wojtek777 at poczta.fm> schrieb im Newsbeitrag 
news:ctqfog$th$1 at korweta.task.gda.pl...
> MMu wrote:
>> some time ago we double stained pma differentiated hl60 cells stained 
>> with fitc (CD36) and tritc (caveoline). the nucleus is stained with dapi.
>> the protein labeled with fitc is a membrane protein (CD36) and thus 
>> should not be seen in the nucleus.
>> the image however, shows green (fitc) wherever dapi (blue) also shows up.
>> we suppose this is some kind of problem with overlapping emmission 
>> wavelengths from dapi and fitc or some kind of similar effect.
>> when looking at the cells under the exictation wavelength for dapi a 
>> shift from blue to green color can be seen over time (usually in less 
>> than 60 seconds).
>> has anyone experienced something like this and knows a solution for this 
>> problem?
> Hi,
> We use fluorescence microscopy a lot. It is definitely problem of filters. 
> To avoid emission overlap from FITC and TRITC other red fluorophores were 
> developed (Texas Red, Cy3). I do not know how to cope with overlap of FITC 
> and DAPI. You can use diffferent green fluorophore, like Oregon Green from 
> Molecular Probes (no affiliation). Contact them, or study their manual to 
> find solution.
> Krzysztof Wasowicz
> Univ Warmia and Mazury,
> Dept Functional Morphol
> Olsztyn, Poland
> wasowicz at uwm.edu.pl 

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