t(11;18)(q21;q21) and aneuploidy are recurrent chromosomal aberrations in mucosa-associated lymphoid tissue (MALT) lymphomas. To investigate their relationship and clinical significance, we developed a two-color fluorescence in situ hybridization (FISH) technique to detect t(11;18) and aneuploidy in nuclei isolated from paraffin-embedded tissue. Thirty-seven MALT lymphomas (all previously evaluated for t(11;18) by reverse transcriptase-polymerase chain reaction), 1 large cell lymphoma (LCL) arising subsequent to MALT lymphoma, and 16 controls were tested by FISH using the t(11;18) probe set and multiple centromeric probes. t(11;18)(q21;q21) was present by FISH in 11 of 12 polymerase chain reaction-positive MALT lymphomas (92%). The LCL and its clonally identical antecedent MALT lymphoma both showed t(11;18). The LCL had trisomy 12, and a small subset of MALT lymphoma cells had trisomy 3 and/or 12. Only one other MALT lymphoma with t(11;18) showed aneuploidy (trisomy 3) in a small clone, whereas 15 of 25 t(11;18)-negative MALT lymphomas (60%) showed trisomy of chromosomes 18 (n = 12), 3 (n = 8), 7 (n = 2), and/or 11 (n = 1). t(11;18) and aneuploidy are primarily mutually exclusive events, suggesting different pathogenetic pathways in the development of MALT lymphomas. Both t(11;18) and aneuploidy were seen disproportionately in lung, and both were associated with recurrent disease.