The Gen-Probe Rapid Diagnostic System for legionellae, which uses 125I-labeled cDNA directed against the rRNAs of legionellae, was evaluated for its ability to detect members of the genus by using clinical specimens which had been frozen at -70 degrees C for 2 to 8 years. Culture and direct immunofluorescence (DFA) results obtained at the time of specimen collection were used to categorize samples. The specimens tested were 112 samples culture positive for legionellae and 230 samples negative on culture and DFA tests. They were tested in a blinded and randomized fashion. Results were expressed in terms of the ratio of counts per minute of the sample to the counts per minute of the provided negative control. A ratio of greater than or equal to 4.0 was picked for optimal specificity. Of the 112 previously positive specimens, 63 (57%) were positive by the probe assay, and of the 230 previously negative samples, 228 (99.1%) were negative. The 51 discrepant specimens were reexamined by culture and DFA testing if adequate amounts remained; this was possible for 34 specimens. On repeat culture, 22 of 33 previously culture-positive samples yielded legionellae and 11 were negative. Ten of the positive repeat cultures yielded two or fewer colonies per plate. One probe-positive but previously culture-negative sample was overgrown by contaminants on repeat culture. Reanalysis of data after exclusion of the 17 unavailable, 11 repeat culture-negative, and 1 unevaluable specimen gave a probe sensitivity of 74% and specificity of 100%. The Gen-Probe test is therefore specific and is of useful sensitivity.