A variety of markers have been used for surface labeling with the scanning electron microscope (1). They are all recognized in the secondary electron imaging (SEI) mode on the basis of their characteristic sizes and shapes. This makes it difficult, however, to recognize unambiguously markers of small size (smaller than 40 nm). Unfortunately, markers of such size are needed if one wishes to minimize steric hindrance phenomena (2) and therefore obtain high efficiency labeling. We have found that the atomic number contrast of the colloidal gold marker, expressed in the backscattered electron imaging (BEI) mode, alleviates this difficulty to a significant extent (3).Human peripheral blood leukocytes, separated by sedimentation in presence of Dextran, were attached to poly-1-lysine pretreated carbon coated grids. The cells were then fixed for 5 min with 0.25% solution of buffered glutaraldehyde, pH 7.2, extensively rinsed with 0.1% glycine in PBS, and incubated for 30 min, at room temperature with an IgM monoclonal antibody (D2) specifically recognizing, a surface antigen of mature granulocytes.
BACKSCATTERED ELECTRON IMAGING OF CRYSTAL MATRIX PROTEIN ON THE SURFACE OF CALCIUM OXALATE CRYSTALS USING COLLOIDAL GOLD
