Elevated　levels　of　intracellular　hydrogen　peroxide　(H2O2)　are　closely　related　to　the　development　of　cancers.　Specific　imaging　of　H2O2　in　tumor　sites　would　be　significant　not　only　for　cancer　diagnosis　but　also　for　gaining　a　deep　understanding　of　the　role　of　H2O2　in　cancer.　However,　traditional　fluorescent　probes　based　only　on　responses　to　overexpression　levels　of　H2O2　in　cancer　cells　are　insufficient　to　distinguish　cancer　cells　from　other　unhealthy　or　healthy　cells　in　complex　biological　systems.　Herein,　we　developed　a　smart,　two-photon　fluorescent　GC-NABP　nanoprobe　with　pH-dependent　surface　charge　conversion　for　tumor-targeted　imaging　of　H2O2.　The　nanoprobe　was　constructed　by　the　self-assembly　of　amphiphilic　GC-NABP,　which　was　synthesized　by　grafting　the　hydrophobic,　H2O2-responsive　and　two-photon　fluorophore,　NABP,　onto　hydrophilic　biopolymer　glycol　chitosan　(GC).　Taking　advantage　of　pH-titratable　amino　groups　on　GC,　the　nanoprobe　had　the　capability　of　surface　charge　conversion　from　negative　at　physiologic　pH　to　positive　in　the　acidic　tumor　microenvironment.　The　positive　charge　of　the　nanoprobe　promoted　electrostatic　interactions　with　cell　membranes,　leading　to　enhanced　cellular　uptake　in　acidic　environment.　Upon　cellular　uptake,　the　high　level　of　H2O2　in　tumor　cells　triggered　boronate　deprotections　of　the　nanoprobe,　generating　a　＇turn-on＇　fluorescence　emission　for　H2O2　imaging.　The　nanoprobe　exhibited　good　sensitivity　and　selectivity　to　H2O2　with　a　detection　limit　down　to　110　nM　in　vitro.　The　results　from　flow　cytometry　and　two-photon　fluorescence　imaging　of　H2O2　in　living　cells　and　tissues　evidenced　the　enhanced　cellular　uptake　and　targeted　imaging　of　intracellular　H2O2　in　acidic　environment.　Compared　to　control　nanoparticles　that　lack　pH　sensitivity,　our　nanoprobe　showed　enhanced　accumulation　in　tumor　sites　and　was　applied　to　targeted　imaging　of　H2O2　in　a　tumor-bearing　mouse　model.　This　work　demonstrates　that　the　nanoprobe　GC-NABP　holds　great　promise　for　tumor-specific　imaging　of　cellular　H2O2,　providing　a　potential　tool　to　explore　the　role　of　H2O2　in　tumor　sites.　This　journal　is　©　The　Royal　Society　of　Chemistry.