Rapid　and　accurate　detection　of　human　epidermal　growth　factor　receptor　2　(HER2)　is　crucial　for　the　early　diagnosis　and　prognosis　of　breast　cancer.　In　this　study,　we　reported　an　iron-manganese　ion　N-doped　carbon　single-atom　catalyst　(FeMn-NCetch/SAC)　bimetallic　peroxidase　mimetic　enzyme　with　abundant　active　sites　etched　by　H2O2　and　further　demonstrated　unique　advantages　of　single-atom　bimetallic　nanozymes　in　generating　hydroxyl　radicals　by　density　functional　theory　(DFT)　calculations.　As　a　proof　of　concept,　a　portable　device-dependent　electrochemical-photothermal　bifunctional　immunoassay　detection　platform　was　designed　to　achieve　reliable　detection　of　HER2.　In　the　enzyme-linked　reaction,　H2O2　was　generated　by　substrate　catalysis　via　secondary　antibody-labeled　glucose　oxidase　(GOx),　while　FeMn-NCetch/SAC　nanozymes　catalyzed　the　decomposition　of　H2O2　to　form　OH*,　which　catalyzed　the　conversion　of　3,3′,5,5′-tetramethylbenzidine　(TMB)　to　ox-TMB.　The　ox-TMB　generation　was　converted　from　the　colorimetric　signals　to　electrical　and　photothermal　signals　by　applied　potential　and　laser　irradiation,　which　could　be　employed　for　the　quantitative　detection　of　HER2.　With　the　help　of　this　bifunctional　detection　technology,　HER2　was　accurately　detected　in　two　ways:　photothermally,　with　a　linear　scope　of　0.01　to　2.0　ng　mL-1　and　a　limit　of　detection　(LOD)　of　7.5　pg　mL-1,　and　electrochemically,　with　a　linear　scope　of　0.01　to　10　ng　mL-1　at　an　LOD　of　3.9　pg　mL-1.　By　successfully　avoiding　environmental　impacts,　the　bifunctional-based　immunosensing　strategy　offers　strong　support　for　accurate　clinical　detection.　©　2024　The　Authors.　Published　by　American　Chemical　Society.