RE: [Histonet] Spleen autofluorescence

From:"Tan, MinHan"

If you think it is autofluorescence due to tissue, which is possible,
view it before adding any antibody.

Actually, I think formalin-fixed tissue tends to have high

I attach here a post which I found on Histonet.

From: "J. A. Kiernan"  


Entering autofluorescence as a search term at brought up 100 hits
all dated in the last 3 years. This is a problem
many people have, and the number and variety of
remedies indicate that there's no perfect solution.
In one recent comparative study, Baschong, Suetterlin
and Laeng (J. Histochem. Cytochem. 49:1565-1571, 2001)
found that staining for 30 min with 0.1% sudan black B 
in 70% alcohol, after immunofluorescent staining,
solved most autofluorescence problems. After staining
the dye was washed off with a jet of a modified Hanks's
solution and then immersed in the same solution for
10 minutes. The authors said this washing was necessary
to avoid formation of a black precipitate.

                            John Kiernan
                            London, Canada
"Hines, Edith M." wrote:
> >         I am having some difficulty overcoming background staining
> > formalin-fixed, paraffin-embedded, human lung tissue sections.  I
> tried blocking
> > the tissues with chromotrope 2R, and with ammonium chloride.  Both
> these
> > methods have not significantly reduced the amount of
autofluorescence that
> I
> > observe.
> >          Any advice that may be able to provide would be greatly
> > appreciated.
> >
> > Thank you for your time,
> > Edie Hines
> > Research Technologist
> > Mayo Clinic Scottsdale
> >
> >

-----Original Message-----
From: Helen Newbery [] 
Sent: Tuesday, May 25, 2004 5:14 AM
Subject: [Histonet] Spleen autofluorescence

Hi histonetters,

Apologies if my query is really basic but I am wanting to do
immunofluorescent antibody staining on mouse spleen sections
(paraffin-embedded) and am seeing a lot of autofluorescence, which I
presume is due to platelets. Is there any way around this problem?

Helen Newbery,
Medical Genetics Section,
Molecular Medicine Centre,
Western General Hospital,
EH4 2XU.
0131 651 1047

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