I used EDTA as a bone decalcifier, and thought the bones were completely decalcified by mechanical manipulation of the bone. On hindsight, they are still very hard. I am going to TRAP stain them and cannot use acid decalcifier, but have never heard of anyone using EDTA as a block surface decalcifier. Does anyone have any information on surface decaling using EDTA? Or could I melt down the blocks and submerge the bones once again into EDTA?
For a bone embedded in a resin (PMMA, epoxy) decalcifiyng with EDTA is a standard procedure. Should also work for fresh bone embedded in paraffin (I believe), but you have not fresh, but partially decalcified bone. It means paraffin may have already infiltrated exposed collagen network and shields hydroxyapatite. In this case you'll need to remove paraffin (melting, soaking/washing in xylene), but I have no idea how it can affect your staining.
First I would try to scrap away the paraffin above the tissue or trim the block as well as possible. So the decalcifier has a chance to reach the hard parts. Then let it swim surface down in EDTA. Watch it after a few minutes. If the tissue becomes whitish and swells out of the block, stop the procedure. This would damage the tissue.
You can try to cut the block again. Sometimes the cause of the hardness is not the insufficient decal, but just the brittleness of collagen after processing. After this "rehydration" of the surface hard blocks may be better to cut.
If this short swimming does not improve cutting and the block looks unaffected you can let it swim for a longer time (and watch).
If all this does not lead to success, melt the block again, and reverse the processing-procedure (xylen - abs. ethanol-96% - 70% - water - EDTA).
I have done trap staining on EDTA-decalcified mouse limbs. As others said, deparaffinize the tissue back till water step, and keep more time in 10%EDTA or 14%EDTA, whatever you have been using. Trap should work on paraffin-sections, Good luck
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