Intracellular pH clamping of human leucocytes: a technique for determination of cellular buffering power and Na+/H+ antiport characteristics.

1. A modified method is described for holding the intracellular pH of leucocytes at different values for prolonged periods of time. 2. Using ammonium sulphate at a final concentration of 8 mmol/l, the buffering power of leucocytes was measured at different internal pH values. 3. A marked dependence of buffering power on internal pH was found. As internal pH was lowered below 7.0, buffering power rose steeply. Above this value, buffering power remained approximately constant. This property may be one factor preventing intracellular acidosis under stress. 4. This method is also suitable for the study of Na+/H+ antiport activity. The addition of external Na+ to cells acid-loaded to an internal pH of 6.7 led to an ethyl isopropyl amiloride-sensitive 1:1 stoichiometric exchange of external Na+ for internal H+.