IL-8 from R&D Systems

From:Mark Elliott

Does anyone have any experience using this antibody from R&D Systems. 
It is a goat anti-human IL-8.  We are trying to stain cryostat cut
sections of guinea pig lung with it but are having problems. 
Specifically we are having problems mostly with background staining.  We
have tried fixing with acetone (10 mins) which seems to give us specific
staing but in our neg control (normal goat serum) we get staining of
some cells as well, especially the airway epithelium.  we have diluted
out our Primary until we get similar staining as in the negative. 
Fixing with 4% paraformaldehyde gets rid of what we think is the
specific staining but the staining on the neg control is still there. We
are using Universal Block from Dako as our blocking reagent and include
1% guinea pig serum and 1% rabbit serum in TBS in all our antibody
dilutions.  Our secondary is biotinylated rabbit anti-goat from Dako at
1/500 dilution and we useDAKO's ABC system using Alkaline Phosphatase
and New Fuchsin as substrate.  Endocenous biotin is not the problem as
we have used ABC before with other antibodies on guinea pig lung and not
had to worry about it.  Any suggestions would be greatly appreciated.


W. Mark Elliott, PhD
Research Associate
UBC-McDonald Research Labs
St. Paul's Hospital
Vancouver, BC

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