The characteristics of [3H]-ouabain binding to human lymphocytes

The kinetic characteristics of [3H]-ouabain binding to human lymphocytes and mixtures of mononucleated cells, and the maximum [3H]-ouabain binding capacities of these cells were studied. The [3H]-ouabain binding was compatible with a single class of receptors with a high affinity for the drug. No signs of positive or negative cooperativity could be demonstrated. In six experiments with pure lymphocyte preparations, the association and dissociation rate constants were 3.08 +/- 0.34 X 10(4)/M/S and 1.58 +/- 0.50 X 10(-4)/S. The dissociation constant derived from equilibrium studies on lymphocytes was 0.68 +/- 0.21 X 10(-8) M, which was identical to that of mononucleated cells. In healthy subjects the maximum [3H]-ouabain binding capacities, which reflect the number of sodium/potassium pump sites were 43154 +/- 8037 molecules/cell (n = 25) in lymphocytes and 75474 +/- 6764 (n = 9) molecules/cell in mixtures of mononucleated cells. Direct determination of the [3H]-ouabain binding capacity of lymphocytes can be performed with acceptable accuracy and precision using 30 ml whole blood. Provided high cell purity, this method may be useful, when studying sodium/potassium pump function in clinical settings.