An improved postembedding technique for ultrastructural studies of lectin binding sites in bone marrow: a critical...

Little is known about the ultrastructural localization of lectin binding sites in human bone marrow tissue. This is probably due to the lack of suitable methods yielding both satisfactory tissue preservation and optimal labelling results. For this reason, we developed a modified postembedding technique for electron-microscopic studies of the glycosylation pattern of haematopoietic cells. Fixation with a 2.5% glutardialdehyde solution was shown to be an important prerequisite and could be even improved by postfixation with tannic acid and uranyl acetate. In particular, embedding with the acrylic resin UnicrylR (Bioacryl) resulted in an optimal ultrastructural preservation of bone marrow tissue. Employment of this hydrophilic resin in combination with a two-step labelling method which included digoxigenin-conjugated Concanavalin A (Con A) followed by ultrasmall anti-digoxigenin-gold and silver amplification, delivered a highly specific staining pattern. Our results with this lectin in different bone marrow cells revealed nuclear and cytoplasmic membranes, rough endoplasmic reticulum as well as granules to display reactivity of varying intensity. These findings underline the validity of our method, for they confirm and extend formerly reported histochemical and biochemical evaluations on the cellular binding sites of Con A.