RNase treatment generally is very useful. But some RNase brands may still 
contain spurious amounts of DNase. DNase activity can be inactivated by the 
following protocol (from SAMBROOK J, FRITSCH EF, MANIATIS T (1989) 
Molecular Cloning: A Laboratory Manual., Vol.3, B17):

dissolve RNase A to 10 mg/ml in 10 mM Tris-HCl pH 7.5, 15 mM NaCl
incubate 15 min at 100 C
cool down slowly (!) (just place in an heat-insulating container, to enable 
renaturation of RNase)
store aliquots at -20 C
avoid prolonged storage at room temperature or in the refrigerator to 
prevent possible renaturation of DNase.

Incubation with PI/RNase (PI: up to 5-10 ug/ml) may also be performed 
overnight at 0-4 C for better equilibration/quantitation.

This protocol may be useful for detection of minor amounts of DNA and for 
quantification (like in flow cytometry experiments).

Guenter


At 15:44 23.01.01 +0200, you wrote:
>RNase digestion?
>(1µg/ml RNase A in PBS, 30 min @ RT)
>Nicely reduces cytosolic staining of PI.
>
>--
>Zsolt Lenkei, MD, PhD
>Laboratoire de Neurobiologie
>ESPCI - CNRS UMR 7637
>10, rue Vauquelin
>75005 Paris, France
>Tel: (33-1) 40 79 51 84
>Fax:  (33-1) 40 79 47 57
>
>
>
> > From: Daniel Harrington <[log in to unmask]>
> > Reply-To: Confocal Microscopy List <[log in to unmask]>
> > Newsgroups: bit.listserv.confocal
> > Date: Sat, 20 Jan 2001 10:48:26 -0600
> > To: [log in to unmask]
> > Subject: Re: Nuclear fluorochrome
> >
> > Does anyone have any suggestions for reducing cytosolic staining with
> > Sytox green?  I'm using it with 3T6 fibroblasts, and find that it
> > stains the nucleus very nicely, but that there's still a fair bit of
> > indiscriminate staining in the remainder of the cell body.  Any tips?
> >
> > --Dan.
> >
> >> Hi Tony,
> >>
> >> Try Sytox Green for fixed cells (exc 488 , green emission). For live
> >> cells have a look of all Syto- carbocyanines. All of them are from
> >> Molecular Probes. They work well if you use at low concentrations.
> >
> > _____________________________________________________________
> > Daniel Harrington
> > E-mail: [log in to unmask]
> > Northwestern University
> > Department of Materials Science and Engineering
> > 2225 N. Campus Dr
> > Evanston, IL 60208-3108
> >
> > Office phone: 847-467-6431            Office: MLSB 3044
> > Lab phone: 847-467-6417               Lab: MLSB 3032
> > 847-467-6416                    MLSB 3026
> >

----------------------------------
Dr. Guenter Giese
MPI fuer Medizinische Forschung
Light Microscopy Facility
Dept. of Biomedical Optics
Jahnstr. 29
D-69120 Heidelberg
Phone (Germany or 0-)6221-486-360
Fax   (Germany or 0-)6221-486-325
e-mail: [log in to unmask]
http://wbmo.mpimf-heidelberg.mpg.de/Biomedizinische.Optik.html
------------------------------------------