FPs - mPlum - spectral separation

[Cameron, Lisa]

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Hello -

I was wondering what people's experience is with the far red FP variants - mPlum
or mRasberry
Roger Tsien's Nat. Methods paper from 2005 says mPlum is better. Have people
successfully been using this FP?
Would be great to hear recommendations/things to consider.
And if one would also want to image eGFP and another FP in between, what would
people recommend for the optimal combination of spectral separation but also
fluor brightness/stability?

I have a core user who is aiming to express all three - I realize this is a
difficult task, just for cell health and transfection, but was hoping for FP
choice recommendations.

Thank you for any feedback -
Feel free to contact me off list.
Lisa


---------------------------------------
Lisa Cameron, Ph.D.
Director of Confocal and Light Microscopy Core
Dana Farber Cancer Institute
Boston, MA 02215
[hidden email]


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[Kurt Thorn]

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We have a couple folks who have successfully used iFP1.4 in the far red
(Cy5) channel on our scopes.  On our four line spinning disk confocal
(405/491/561/640) we have successfully done 4-color imaging in mammalian
cells using mTagBFP / EGFP / mCherry / iFP1.4.  Probably there are
better combinations out there (iRFP is supposed to be better than
iFP1.4) but this one works well.

Kurt

On 11/2/2011 3:10 PM, Cameron, Lisa wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> *****
>
> Hello -
>
> I was wondering what people's experience is with the far red FP variants - mPlum
> or mRasberry
> Roger Tsien's Nat. Methods paper from 2005 says mPlum is better. Have people
> successfully been using this FP?
> Would be great to hear recommendations/things to consider.
> And if one would also want to image eGFP and another FP in between, what would
> people recommend for the optimal combination of spectral separation but also
> fluor brightness/stability?
>
> I have a core user who is aiming to express all three - I realize this is a
> difficult task, just for cell health and transfection, but was hoping for FP
> choice recommendations.
>
> Thank you for any feedback -
> Feel free to contact me off list.
> Lisa
>
>
> ---------------------------------------
> Lisa Cameron, Ph.D.
> Director of Confocal and Light Microscopy Core
> Dana Farber Cancer Institute
> Boston, MA 02215
> [hidden email]
>
>
> The information in this e-mail is intended only for the person to whom it is
> addressed. If you believe this e-mail was sent to you in error and the e-mail
> contains patient information, please contact the Partners Compliance HelpLine at
> http://www.partners.org/complianceline . If the e-mail was sent to you in error
> but does not contain patient information, please contact the sender and properly
> dispose of the e-mail.
>

[Peter Humphreys]

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I have a couple of users with uGFP, mKO and mcherry.
No problems on our Leica SP5. The mKO is exceptionally bright in their
samples.

Peter
CSCR, Cambridge


  On 03/11/11 16:51, Kurt Thorn wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> *****
>
> We have a couple folks who have successfully used iFP1.4 in the far
> red (Cy5) channel on our scopes.  On our four line spinning disk
> confocal (405/491/561/640) we have successfully done 4-color imaging
> in mammalian cells using mTagBFP / EGFP / mCherry / iFP1.4.  Probably
> there are better combinations out there (iRFP is supposed to be better
> than iFP1.4) but this one works well.
>
> Kurt
>
> On 11/2/2011 3:10 PM, Cameron, Lisa wrote:
>> *****
>> To join, leave or search the confocal microscopy listserv, go to:
>> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>> *****
>>
>> Hello -
>>
>> I was wondering what people's experience is with the far red FP
>> variants - mPlum
>> or mRasberry
>> Roger Tsien's Nat. Methods paper from 2005 says mPlum is better. Have
>> people
>> successfully been using this FP?
>> Would be great to hear recommendations/things to consider.
>> And if one would also want to image eGFP and another FP in between,
>> what would
>> people recommend for the optimal combination of spectral separation
>> but also
>> fluor brightness/stability?
>>
>> I have a core user who is aiming to express all three - I realize
>> this is a
>> difficult task, just for cell health and transfection, but was hoping
>> for FP
>> choice recommendations.
>>
>> Thank you for any feedback -
>> Feel free to contact me off list.
>> Lisa
>>
>>
>> ---------------------------------------
>> Lisa Cameron, Ph.D.
>> Director of Confocal and Light Microscopy Core
>> Dana Farber Cancer Institute
>> Boston, MA 02215
>> [hidden email]
>>
>>
>> The information in this e-mail is intended only for the person to
>> whom it is
>> addressed. If you believe this e-mail was sent to you in error and
>> the e-mail
>> contains patient information, please contact the Partners Compliance
>> HelpLine at
>> http://www.partners.org/complianceline . If the e-mail was sent to
>> you in error
>> but does not contain patient information, please contact the sender
>> and properly
>> dispose of the e-mail.
>>