SummarySpecific mRNAs coding for a 63 kDa sperm membrane protein (63-SMP) were localised in Strongylocentrotus purpuratus testis sections using in situ hybridisation techniques. ³⁵S-labelled antisense RNA probes transcribed from a 766 base pair fragment of the gene coding for the 63-SMP hybridised to all spermatogenic cells in the basal germinal epithelia of testicular acini, except the most peripherally located (least differentiated) spermatogonia. No hybridisation to the luminally located mature spermatozoa or somatic cells of the testis was observed. Using monoclonal antibody J17/30 and indirect immunofluorescence techniques, the 63-SMP was localised to the same subset of spermatogenic cells that contain the 63-SMP mRNA, suggesting that expression of this gene is transcriptionally controlled. In combination with previous studies on the expression of sperm histones and sperm bindin, these results show that multiple, perhaps sequential, classes of gene activity contribute to the differentiation of sea urchin sperm.