Hapten-tagged Plasma Proteins As Immunocytochemical Probes for the Study of Vascular Permeability

Bovine serum albumin and transferrin were covalently coupled with fluorescein isothiocyanate and digoxigenin, respectively, and intravenously co-injected in equal amounts in mouse. The derivation of the two proteins induces minor alterations of their physicochemical properties as well as of their physiological functions. The two tracers were revealed within vascular and extravascular compartments of diaphragm by quantitative postembedding immunocytochemistry, using antibodies against each of the haptens in conjunction with the protein AG-gold complexes. The influence of different fixatives and embedding protocols on the immunodetectability of the hapten-tagged proteins was assessed. Both resist reasonably well to osmication and embedding in Epon. None of the haptens reacted with the heterologous antibody. At 30 minutes after injection, the tracers were detected in blood plasma, interstitium, and endothelial plasmalemmal vesicles. The presence of both proteins within the interendothelial clefts was inconspicuous. The ratios between the labeling densities found over endothelium, interstitial space, and vascular lumen were similar for both tracers. This suggests that the endothelium of mouse diaphragm capillaries might exhibit comparable permeabilities towards serum albumin and transferrin which are similar in size and charge. The study shows that hapten-tagged polypeptides are close to the corresponding native macromolecules, and represent interesting tools for the morphological study of dynamic processes such as transcytosis.