BACKGROUND: Saponin is a mild detergent commonly used to permeabilize cells prior to immunofluorescence labeling of intracellular proteins. It has previously been used to that effect in Dictyostelium discoideum amoebae. FINDINGS: We show that saponin, contrary to Triton X-100 or alcohol, permeabilizes at best incompletely membranes of Dictyostelium. In cells exposed to osmotic stress, almost complete resistance to saponin permeabilization was observed. CONCLUSIONS: Saponin should be used with special care to permeabilize Dictyostelium membranes. This unsusual property is presumably linked to the specific sterol composition of Dictyostelium membranes. It may also represent an adaptation of Dictyostelium to harsh conditions or to natural compounds encountered in its natural environment.