Vicinal glycol-staining identifies secondary granules in human normal and Chédiak-Higashi neutrophils

Neutrophil secondary granules contain large amounts of glycoprotein. We evaluated periodate-thiocarbohydrazide-silver proteinate (PA-TCH-SP) staining of these granules after alpha-amylase digestion to assess their content of vicinal glycol-containing glycoconjugates and the usefulness of this stain as a positive stain for secondary granules. Using this method, early stages of secondary granule genesis were observed prior to completion of primary granule genesis in myelocytes. Immature secondary granules appeared round to ovoid, but irregular in outline and demonstrated strong staining of the limiting membrane and matrix material. In mature granules, matrix staining was unaltered; however, membrane staining was decreased. Some immature primary granules in promyelocytes demonstrated strong PA-TCH-SP reactivity which was masked in mature primary granules of band and segmented neutrophils. The Golgi apparatus showed progressively increasing PA-TCH-SP reactivity toward its mature surface which was often convex in promyelocytes and myelocytes and concave in segmented neutrophils. The Chédiak-Higashi secondary granules were cytochemically and morphologically similar to those of normal individuals and were not statistically decreased in number when compared to controls. They were only rarely observed contacting or fusing with giant granules which had consumed all primary granules leaving an easily detected population of secondary granules. Thus the alpha-amylase-PA-TCH-SP method demonstrates a large amount of unmasked vicinal glycol-containing glycoconjugates in neutrophil secondary granules, which allows their differentiation from primary granules.