APPLICATION OF X-RAY-MICROANALYSIS TO STUDY OF THE EXPRESSION OF ENDOTHELIAL ADHESION MOLECULES ON HUMAN UMBILICAL VEIN ENDOTHELIAL-CELLS IN-VITRO

A semi-quantitative procedure is described, which allows the evaluatio n of expression levels of endothelial adhesion molecules on cultured h uman umbilical vein endothelial cells (HUVEC) using energy dispersive X-ray microanalysis (EDX). As a model two adhesion molecules, E-select ion (CD62E; ELAM-1/endothelial leukocyte adhesion molecule-1) and ICAM -1 (intercellular adhesion molecule-1; CD54), were localized by the us e of the silver-enhancement colloidal gold method after stimulation of HUVEC with endotoxin lipopolysaccharide (LPS), tumour necrosis factor (TNF) or a phorbol ester (PMA). The analysis was performed in a scann ing electron microscope (SEM) at an accelerating voltage of 15 kV with scanned areas of 200x400 mu m. The semi-quantitative data indicated t hat in LPS-treated groups both adhesion molecules were expressed at a significantly higher level than in all other groups (P<0.01). In addit ion, after a 4 h treatment the expression levels of E-selectin in all groups were higher compared to ICAM-1. The experimental data from X-ra y microanalysis were compared with data obtained from an enzyme-linked immunosorbent assay (ELISA) and similar Values were found for both ty pes of preparation. This microanalytical method is relatively simple a nd seems to be suitable for immunogold labelling studies on different types of endothelial cells in vitro.