The separation of different cell classes from lymphoid organs. The effect of pH on the buoyant density of lymphocytes and erythrocytes.

Abstract

SummaryThe buoyant density of erythrocytes suspended in serum, or in iso‐osmotic. media at neutral pH, is around 1·108 g/cm3. Lowering the pH of the medium, from pH 7 to pH 5. without change of osmolarity, causes about a 50% increase in volume and a decrease it density of about 0·02 g/cm3. It was established that this was due to the pH change itself, und not to changes in cation concentration. The basis for the swelling is an accumulation of anions in the cell as the pH is lowered, in accordance with the Donnan equilibrium across the membrane.Lymphocytes do not display pH induced volume changes, nor do they change in density with pH when tested by a phthalate ester non‐aqueous density separation procedure. This is as expected from the relative ion impermeability of the membrane. However, lymphoid cell buoyant density does change with pH if separation is accomplished in continuous, iso‐osmotic gradients of albumin. This effect is reversible, and amounts to a 0·01 g/cm3 lower density at pH 5·1 than at pH 7·2. The buoyant density value at pH 5·1 in albumin gradients is close to the density of cells in scrum, as estimated by the non‐aqueous separation procedure.The basis for the anomalous behaviour of lymphocytes in continuous. aqueous density gradients is discussed ill relation to the density analysis of lymphoid cells and the pH‐shift technique for the separation of erythroid from lymphoid cells.