Two distinct CD3 homologue genes, $CD3gamma/delta;and;CDvarepsilon$, were isolated from a olive flounder leukocyte cDNA library and a BAC library. $CD3gamma/delta$ consisted of 961 bp encoding 178 amino acid residues, and $CD3varepsilon$ consisted of 1006 bp encoding 164 amino acid residues. When compared with other known CD3 peptide sequences, the most conserved region of the two olive flounder CD3 chain peptides are the cytoplasmic domain and the least conserved are the extracellular domain. A phylogenetic analysis based on the deduced amino acid sequence grouped the two olive flounder CD3 sequences with $CD3varepsilon$ and $CD3gamma/delta$, respectively. The olive flounder CD3 cluster (consisting of $CD3varepsilon;and;CD3gamma/delta$) spans only 10.4 kb. The $CD3varepsilon;and;CD3gamma/delta$ genes are oppositely transcribed only 3.8 kb apart. Both olive flounder CD3 genes have five exons. The two olive flounder CD3 genes were predominantly expressed in PBLs, kidney, spleen, and gills.