Quantitative assay for lectin-induced cytoagglutination by means of an electronic particle-counting technique

A method employing an electronic particle-counting technique was used to quantify lectin-induced agglutination of human granulocytes and lymphocytes with either concanavalin A or wheat germ agglutinin. The number and mean volume of single cells and aggregates in the presence of increasing concentrations of lectin were computed from 95% confidence intervals. Agglutination depended on both the number of free cells and the number and size of the cell clusters. Changes in these two variables were mutually independent of one another, and both were simultaneously determined. An index of agglutination that takes the effect of these two variables into account was defined as (formula: see text) VA equals mean volume of cell aggregates, NS equals number of single cells, NA equals number of cell aggregates, VS equals mean volume of single cells, rb equals r at a given lectin concentration, and ra equals r in the absence of lectin. For any combination of lectin and cell type, the agglutination curve, as described by zeta, consisted of two components: a) a flat region in which zeta remained constant with increasing lectin concentrations and b) a region in which zeta increased linearly as a function of the logarithm of lectin concentration. The shapes of these curves offered two parameters for quantitative comparison of agglutinability: 1)threshold concentration, defined as the minimum concentration of lectin (microgram/ml) required to bring about a measurable rise in zeta and 2) the concentration gradient, defined as the change in zeta for an increase of one log unit in the concentration of the lectin in the range beyond the threshold concentration. This method offers a high degree of quantification and provides reliable information that can be meaningfully correlated with cell surface characteristics.