An improved method for identifying 8-O-acetyl-N-acetylneuraminic acid.

The periodic acid/thionin-Schiff/potassium hydroxide/periodic acid/fuchsin-Schiff sequence developed by Culling et al. frequently causes damage to sections and gives inconsistent results because of insufficient primary oxidation and difficulties in making the thionin-Schiff reagent. These disadvantages have been largely eliminated by more thorough primary oxidation and by replacing the original thionin-Schiff with a new cold thionin-Schiff. The effect of alkaline hydrolysis on thionin-aldehyde complexes was also studied and the reduction of color caused by this treatment was restored by a second thionin-Schiff reaction. The new sequence gives consistent results and imparts greater color to the thionin-Schiff reaction.