The fruitfly, Drosophila melanogaster, has three proPO genes (DoxA1, CG8193 and CG2952). DoxA1 has been shown to encode proPO A1, one of the two proPO isoforms (A1 and A3). However, which of CG8193 or CG2952 encodes proPO A3 has so far remained elusive. In Northern analysis, CG8193 expression was strong during the larval stage, yet expression of CG2952 was not detected at any stage. Immunoblot analyses with specific antibodies detected CG8193 in the larval hemolymph at the mobility of the endogenous proPO A3, though no signal for CG2952. These results indicate that the expression of CG2952 is very low and that CG8193 is the gene that encodes proPO A3. Processing of A1 and A3 isoforms in adult homogenate and activity of recombinant proPOs were also investigated.