Reactive changes in free intracellular zinc cation concentration ([Zn²⁺]_i) were monitored, using the fluorescent probe Zinquin, in human lymphoma cells exposed to the DNA-damaging agent VP-16. Two-photon excitation microscopy showed that Zinquin-Zn²⁺ forms complexes in cytoplasmic vesicles. [Zn²⁺]_i increased in both p53^wt (wild type) and p53^mut (mutant) cells after exposure to low drug doses. In p53^mut cells noncompetent for DNA damage-induced apoptosis, elevated [Zn²⁺]_i was maintained at higher drug doses, unlike competent p53^wt cells that showed a collapse of the transient before apoptosis. In p53^wt cells, the [Zn²⁺]_i rise paralleled an increase in p53 and bax-to-bcl-2 ratio but preceded an increase in p21^WAF1, active cell cycle arrest in G₂, or nuclear fragmentation. Reducing [Zn²⁺]_i, using N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine, caused rapid apoptosis in both p53^wt and p53^mut cells, although cotreatment with VP-16 exacerbated apoptosis only in p53^wt cells. This may reflect changed thresholds for proapoptotic caspase-3 activation in competent cells. We conclude that the DNA damage-induced transient is p53-independent up to a damage threshold, beyond which competent cells reduce [Zn²⁺]_i before apoptosis. Early stress responses in p53^wt cells take place in an environment of enhanced Zn²⁺ availability.