Posted by
Michael Weber-4 on
URL: http://confocal-microscopy-list.275.s1.nabble.com/CFP-YFP-tp590300p590306.html
Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocalHi Park,
based on your post, I assume you want to use a confocal, since you are
talking about channels and excitation wavelength?! Which type of system
are you using?
In general, I can not recommend to simultaneously acquire CFP and YFP,
since there is quite some overlapping between the emission of those both
dyes (see
http://www.mcb.arizona.edu/ipc/fret/index.html). Sequential
detection would be the way to go. You can do this line-by-line with a
confocal, or with dual dichroic plus fast emission filter wheel on a
camera-based system, if you would like to follow rapid events. Common
excitation wavelengths for CFP are 405/440/458, for YFP 514nm. The optimal
filter setup is based on your system (i.e. dichroic performance).
Fluorchrome databases like the one I mentioned above are very useful for
finding the best combination.
cheers,
Michael
> Search the CONFOCAL archive at
>
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal>
> Another important question is the temporal and spatial resolution you
> require. Do you need to look at organelles in one yeast moving "real
> time" or fields of COS, Hela or fibroblasts over many hours?
>
>
>
>> Search the CONFOCAL archive at
>>
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal>>
>> Ok, do you want to do confocal or two photon?
>> Can you afford an off-the-shelf system, or are you building your own
>> from scratch, or do you have an existing system you could modify?
>>
>> Craig
>>
>>
>> On 9/21/07, Lee, Park Joo <
[hidden email]> wrote:
>>> Search the CONFOCAL archive at
>>>
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal>>>
>>> Hi Everyone,
>>>
>>> Please help. What could be a good experimental setup to do live cell
>>> imaging
>>> using CFP and YFP on two separate channels such excitation wavelength
>>> and
>>> emission filters?
>>>
>>> Park
>>>
>>
>
>
> _________________________________________
> Michael Cammer
http://www.aecom.yu.edu/aif/