The aim of this work was to identify a new protein that discriminated CD8 from CD4 and CD19 lymphocytes. Proteins were separated by high-resolution two-dimensional electrophoresis. After silver staining, the gel images were captured with a laser densitometer, and studied with a dedicated software. This study confirmed the presence of two spots that appeared to be preferentially associated with CD8 lymphocytes, and mass spectrometry analyzes (liquid chromatography-tandem mass spectrometry, LC-MS/MS) identified six peptides for one spot and four for the other. The peptide sequences corresponded to an unknown protein that we named swiprosin 1 (Swiss-Prot Q96C19). Molecular analysis (reverse transcriptase-polymerase chain reaction, RT-PCR) and Northern blots confirmed that the gene expression was increased in purified populations of CD8 lymphocytes, when compared to CD19 and CD4 lymphocytes. Database mining revealed that swiprosin 1 contains two potential EF-hand domains, and therefore may have a role in calcium signaling. Its predominant presence in CD8 lymphocytes suggests that it may be involved in functions that are important for cytotoxic lymphocytes.