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We have tried the Flash system and stopped using it due to different 
reasons: It accumulated in mitochondria, and we could only reverse this by 
using EDT or beta-mercaptoethanol, these treatments and the effects on 
mitochondria were not compatible with long live cell imaging. The specific 
staining we got was OK in terms of localization but very prone to 
photobleaching and the signal to noise was clearly inferior to GFP. 
Furthermore, we never succeeded in fixing the staining for EM, like 
suggested in Tsiens publications.
Most of this is in press (Histochemistry and cell biology DOI 
10.1007/s00418-005-0136-3).
Regards
Matthias
----------------------
Matthias F. Langhorst

Developmental Neurobiology Group
University of Konstanz
Universitaetsstr. 10
D-78457 Konstanz
Germany

Tel.: 0049-7531-882128 (office)
    0049-7531-882135 (lab)
Fax 0049-7531-883894

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----- Original Message ----- 
From: "David Knecht" <[log in to unmask]>
To: <[log in to unmask]>
Sent: Tuesday, May 09, 2006 11:57 PM
Subject: Flash tag


> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> I wondered if anyone out there has had any experience with Tsien's  (now 
> INvitrogen) Flash tag system.  We made our first construct  recently 
> containing the tetracysteine motif, but got no fluorescence  at all with 
> the biarsenical compound.  We are checking expression  now. The absence of 
> much literature of this outside of the Tsien lab  has me concerned that 
> there are issues with making it work.
> Dr. David Knecht
> Department of Molecular and Cell Biology
> U-3125
> 91 N. Eagleville Rd.
> University of Connecticut
> Storrs, CT 06269
> 860-486-2200
> 860-486-4331 (fax)