A family of synapse-associated protein(SAPs) has recently emerged as a central player in the molecular organization of synapses. This work is to analyze the epitope of a novel Homo sapiens synapse associated-protein(FRG4) and synthesize polyclonal antibody, then research FRG4 protein expression in smooth muscle cells and endothelial cells. FRG4 full-length sequence was obtained by PCR from human fetal liver library. Its epitope, motifs and the second structure of amino acids encoded were detected by bioinformatics technique. By solid-phase peptide synthesis method to synthesize FRG4 peptides, then peptides were immunized to rabbits; by immunohistochemistry to detect the expression of FRG4 in vascular smooth muscle cells and endothelial cells. The 13-peptides PKLVKEEVFWRNY was selected by bioinformatic analysis to synthesize rabbit anti-human FRG4 polyclonal antibody. Antibody purity was 82.79% and antibody dilution was 1∶16 000 detected by ELISA. The antibody has a good reaction and speciality in Western blot, it was mainly expressed in the cytoplasm of smooth muscle cells and endothelial cells by immunohistochemistry. The results suggested a novel Homo sapiens synapse associated protein (FRG4) antibody was synthesized successfully. FRG4 expressed mainly in the cytoplasm of vascular smooth muscle cells and endothelial cells.