http://confocal-microscopy-list.275.s1.nabble.com/CFP-YFP-tp590300p590301.html
because you cannot excite the dyes one after each other. And on the
> Thanks Mike. I was thinking about doing it either on confocal or on twp photon. The sequential detection you mentioned sounds very interesting. Maybe I should try it out.
>
> Park
>
> -------------- Original message ----------------------
> From: Michael Weber <
[hidden email]>
>> Search the CONFOCAL archive at
>>
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal>>
>> Hi 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/