We have investigated the internalization to low pH intracellular compartments of transferrin (Tfn), diphtheria toxin (DT) and of anti-cell surface antibodies (MAb) by a cytofluorometric assay based on low pH quenching of fluorescein (FITC) emission. FITC-labelled Tfn, anti-CD3, anti-CD5 and anti-Thy 1.2 MAb internalization resulted in a progressively lower FITC quenching effect. Following internalization, a distinction could be made between molecules that enter low pH compartments without undergoing intracellular degradation (e.g., Tfn, anti-CD3 MAb) and molecules that are internalized through low pH organelles and are then degraded within the cell (e.g., DT). A strict correlation was observed between quenching of internalized FITC-protein fluorescent emission and the cytotoxic activity of DT-based immunotoxins (IT).