Concanavalin-A-mediated thymocyte agglutination: a model for a quantitative study of cell adhesion

Abstract

This report describes a quantitative study of the agglutination of rat thymocytes with concanavalin A (ConA). The probability that two ConA-coated cells remain bound after centrifugation was determined over a wide range of lectin concentrations. The minimal force required to separate agglutinated cells and the number of ConA molecules bound per cell were measured in similar experimental conditions. Agglutinated cells were examined by electron microscopy to estimate the area of membrane involved in adhesion. The dependence of agglutination on cell metabolism was studied: cold (4 degrees C), sodium azide (15 mM) and cytochalasin B (10 micrograms/ml) inhibited thymocyte adhesion. The importance of lateral movements of ConA molecules was assayed by measuring the adhesion of ConA-coated glutaraldehyde-fixed thymocytes to untreated cells: substantial binding occurred, but at a reduced level relative to untreated cells. A mathematical analysis of experimental data allowed the following conclusions. (1) At least 10(3) ConA bonds were involved in cross-linking two bound cells, which required the lectin molecules to be concentrated in the binding area, at least when low ConA concentrations (0.5 microgram/ml or less) were used. (2) The dependence of the binding probability on lectin concentration was fairly linear when the latter was small, which implied that the limiting step in cell-cell adhesion was the formation of a bond between a single ConA molecule and a ligand on the other cell. (3) The mean intercellular-contact time for the formation of this first bond was about 10 S for high concentrations of ligand (8 micrograms/ml). It was possible to fit the above data into a physically consistent quantitative model of cell adhesion.