[Histonet] Quenching auto fluorescence in human tissue

[Daniela Bodemer]

Hi all,

 

I have been experiencing trouble with my secondary antibody when doing
my immuno with human colon tissue.

I use to use FITC 488 as my secondary antibody and changed over to
Alexa488 due to the FITC's manufacturing stop.

The auto fluorescence of the human tissue is quenched with Chicago Blue
solution and the slides are mounted with Prolong Gold antifade reagent
to extend the life of the immuno slides.

The trouble is, after a few days the fluorofors seem to 'detach' and
start floating around, not allowing reliable validation of the results.
This also happen if I use Mowiol as a mounting medium.

Is the mounting medium reacting with Chicago Blue? Is there any
alternative to quench die auto fluorescence of the tissue?

 

Thanks for your ideas.

 

Daniela Bodemer 

Research Assistant

Surgical Research, Infection and Immunity

 

Murdoch Childrens Research Institute

The Royal Children's Hospital

Flemington Road Parkville Victoria 3052 Australia 

T 03 9936 6676     T (03 9936 6794)     

E daniela.bodemer <@t> mcri.edu.au <mailto:firstname.surname <@t> mcri.edu.au> 

www.mcri.edu.au <http://www.mcri.edu.au/> 

 

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