A new method for the histochemical demonstration of O-acyl sugars in human colonic epithelial glycoproteins

A new general method has been developed for the specific histochemical identification of O-acyl sugars in any epithelial glycoprotein. These sugars include hexose, 6-deoxyhexose and N-acetylhexosamine with an ester substituent(s) located on a potential vicinal diol(s). In the procedure reported [the periodic acid-borohydride reduction-saponification-selective periodate oxidation-borohydride reduction-periodic acid-Schiff (PA-Bh-KOH-PA*-Bh-PAS) method] the initial PA-Bh treatment renders vicinal diols located on either sialic acid or neutral sugars PAS unreactive. In the subsequent steps ester substituents are removed from both O-acyl sugars and O-acyl sialic acids by saponification (KOH), sialic acid vicinal diols are selectively removed by the PA*-Bh sequence and O-acyl sugars are stained with the PAS technique. This method has the advantage that the results are obtained with a single section and the results are either positive or negative. Consequently, it is superior to the three indirect methods investigated because it does not require an observer to compare the intensity or the shade of the staining obtained with serial sections. Using the PA-Bh-KOH-PA*-Bh-PAS method we have demonstrated, for the first time, that O-acyl sugars occur in the epithelial goblet cell glycoproteins of adult human colon. The effect of the presence of O-acyl sugars on the interpretation of a number of other methods for the histochemical investigation of glycoproteins is discussed. It is recommended that the results obtained with the PA-Bh-KOH-PA*-Bh-PAS method be evaluated before histochemical procedures for the investigation of neutral sugars and O-acyl sialic acids are selected.