Surface-enhanced　Raman　scattering　(SERS)　has　great　potential　in　biological　analysis　due　to　its　specificity,　sensitivity,　and　non-invasive　nature.　However,　effectively　extracting　Raman　information　and　avoiding　spectral　overlapping　from　biological　background　interference　remain　major　challenges.　In　this　study,　we　developed　a　background-free　SERS　nanosensor　consisting　of　gold　nanobipyramids　(Au　NBPs)　core-Prussian　blue　(PB)　shell　(Au　NBPs@PB),　for　endogenous　H2S　detection.　The　PB　shell　degraded　quickly　upon　contact　with　endogenous　H2S,　generating　a　unique　Raman　signal　response　in　the　Raman　silent　region　(1800-2800　cm-1).　By　taking　advantage　of　the　high　SERS-activity　of　Au　NBPs　and　H2S-triggered　spectral　changes　of　PB,　these　SERS　nanosensors　effectively　minimize　potential　biological　interferences.　The　nanosensor　exhibits　a　detection　range　of　2.0　μM　to　250　μM　and　a　limit　of　detection　(LOD)　of　0.34　μM,　with　good　reproducibility　and　minimal　interference.　We　successfully　applied　this　background-free　SERS　platform　to　monitor　endogenous　H2S　concentrations　in　human　serum　samples　with　satisfied　results.　©　2024　American　Chemical　Society.