Some reports have indicated that trisomy 3 represents a characteristic chromosomal abnormality found in lymphomas arising in mucosa-associated lymphoid tissues (MALT)/extranodal marginal zone B-cells (MZBC). Traditional cytogenetic analysis of metaphase preparations is cumbersome and not always possible, especially in those situations in which the diagnosis in not suspected before a biopsy. Our aim is to use a relatively simple method to evaluate trisomy 3 in paraffin-embedded, formalin-fixed tissue, using fluorescence in situ hybridization (FISH) on intact tissue sections. Formalin-fixed, paraffin-embedded archival tissues from 30 cases (27 lymphoma and 3 chronic gastritis cases) were hybridized with a chromosome 3 specific alpha-satellite probe (ONCOR, Gaithersburg, MD). Three of four cases of gastric MZBC/MALT lymphoma revealed trisomy 3. Ten cases of lymphoma of possible or probable MZBC origin were examined, and four revealed trisomy 3. Five of 13 non-MZBC lymphomas revealed trisomy 3. None of the chronic gastritis cases nor normal tonsil cases revealed trisomy 3. Our results, using a different methodological approach, confirm the findings of others that trisomy 3 is an abnormality found in a significant proportion of lymphomas of MZBC origin. Our approach also makes possible interphase cytogenetic analysis (by FISH) of routinely processed formalin-fixed, paraffin-embedded tissues, without the need to disaggregate cells. It thus may facilitate genetic analysis on specimens previously deemed unsuitable for such analysis, particularly when tissue quantity is limited.