Counterstain for NBT/BCIP


Hi, we are doing some ISH with digoxigenin-labelled probes and an 
anti-DIG-alkaline phosphatase conjugated antibody, on paraffin-embedded 
sections. However, we are having trouble to find the perfect 
counterstain after detection of AP activity with NBT/BCIP (very dark 
bluyish/purple). It would be perfect to use methyl green because then 
results might be recorded fine in B&W photos using a green filter. 
However, in our hands the sections do not get the stain even after 10 or 
more minutes on a warm plate (is this right or are we missing 
something?). We have also tried nuclear fast red and it is OK, but 
contrast is not too good in slides with heavy signal (overall 
pink/purple background). In the past I also used bismark brown Y and it 
gave good contrast, but the staining is very uniform and does not give 
much information about the tissue or the cells present (no nuclear or 
diferential staining). So the question is, any idea on a better 
counterstain or on how to use methyl green in this kind of material?. 
Thanks in advance,

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