Ca2+ influx was measured into human erythrocytes in which efflux was blocked by either introduction of an intracellular Ca2+ chelator, introduction of the Ca2+ chelator followed by ATP depletion, or depletion of the Ca2+ pump cofactors ATP and Mg2+. The Ca2+ influx under all three conditions was 14-20 mumol . 1 cells-1 . h-1, which is an order of magnitude higher than the influx previously reported for cells depleted of either ATP or Mg2+ separately. The difference between the two values was explained by the finding of substantial Ca2+ efflux from the Ca2+-loaded ATP-depleted cells, whereas this efflux was insignificant from cells loaded with quin 2 and then ATP depleted. Under these latter conditions Ca2+ influx estimates the unidirectional permeability to this cation. Studies using this technique showed that Ca2+ influx was the same in media of isotonic sodium, potassium, lithium, choline, or magnesium chlorides. Moreover the dependence of Ca2+ influx on external Ca2+ concentration was well described by the sum of saturable and nonsaturable (linear) components.