RE: GFP

   Our experinece is that peripheral blood mononuclear cells from GFP transgenic mice are resistant to fixation with 1% paraformaldehyde. Fixed cells have been found to fluoresce for at least 2 weeks (may be longer) when kept in the dark in the refrigerator. The fluorescence intensity of fixed cells from different individual GFP mouse does not differ when analyzed within the same day. However, after long-term storage (>2 weeks) in 1% PF  resulted in a small drop in % GFP-expressing cells in some but not all GFP transgenic mice. I guess this may vary among various transgenic mice expressing GFP under different promoters as well.

Sundararajan Jayaraman, Ph.D.
Associate Professor
Dept. of Surgery
University of Illinois at Chicago
909 South Wolcott Avenue
COBRB Room 8113
Chicago, IL 60612
Tel: 312-355-5133
Fax: 312-996-6415




-----Original Message-----
From: Gerstein, Rachel [mailto:Rachel.Gerstein@umassmed.edu]
Sent: Fri 12/8/2006 3:10 PM
To: cyto-inbox
Subject: RE: GFP
 
PFA fixation dampens GFP signal, although fixation can be adjusted to still get some detection.  This has been discussed many times on this list; see www.cyto.purdue.edu/hmarchiv/index.htm

=======================================================
Rachel M. Gerstein, Ph.D.
Associate Professor
Department of Molecular Genetics and Microbiology
Graduate Program in Immunology/Virology
University of Massachusetts Medical School
55 Lake Avenue North
Worcester, MA 01655-0002
(508) 856-1044
(508) 856-5920 (FAX)


> ----------
> From: 	Ibtissam Abdul-Jabbar
> Sent: 	Thursday, December 7, 2006 10:31 PM
> To:	Cytometry Mailing List
> Subject:	FW: GFP
> 
> Dear Flowers would you please answer Ray on his question { whether detection of eGFP by FACS is enhanced or hindered by paraformaldehyde fixation of cells prior to cytometry?}
> 
>  
> 
> Many thanks
> 
> iaj
> 
>  
> 
>   _____  
> 
> From: Ray Steptoe 
> Sent: Thursday, 7 December 2006 4:10 PM
> To: Ibtissam Abdul-Jabbar
> Subject: GFP
> 
>  
> 
> Hi Ibtissam,
> 
>  
> 
> Would it be possible to ask the flow users list whether detection of eGFP by FACS is enhanced or hindered by paraformaldehyde fixation of cells prior to cytometry?
> 
>  
> 
> Many thanks
> 
> Ray
> 
>  
> 
> -----------------------------------------------------------------
> 
> Raymond J. Steptoe  PhD
> 
> The Centre for Immunology & Cancer Research 
> 
> The University of Queensland 
> 
> Princess Alexandra Hospital 
> 
> Level 4, R Wing, Building 1
> 
> Woolloongabba,  QLD 4102 
> 
> AUSTRALIA
> 
> Telephone: + 61 7 3240 5393	Facsimile: + 61 7 3240 5946 
> 
> Internet: http://www.cicr.uq.edu.au 
> 
>  
> 
>  
> 
> 


**************************EMAIL DISCLAIMER***************************

This email and any files transmitted with it may be confidential and are
intended solely for the use of the individual or entity to whom they are
addressed.  If you are not the intended recipient or the individual
responsible for delivering the e-mail to the intended recipient, any 
disclosure, copying, distribution or any action taken or omitted to be 
taken in reliance on it, is strictly prohibited.  If you have received this
e-mail in error, please delete it and notify the sender or contact Health 
Information Management 312.413.4947.