On-site　quick　detection　of　acetylcholinesterase　(AChE)　activity　is　of　great　significance　for　diagnosing　nervous　diseases,　screening　drugs,　and　monitoring　poisoning.　Herein,　we　developed　a　system　for　sensitively　regulating　gold　nanobipyramid　(AuNBP)　morphology　on　a　paper-based　analytical　device　(PAD)　via　an　Au+-disproportionation　system,　and　further　developed　a　high-resolution　paper-based　multicolor　sensor　(PMS)　for　both　on-site　visual　detection　and　laboratory　precision　quantification　of　AChE　activity　using　the　system　as　a　signal　reporter　and　a　dual-enzyme　strategy　as　a　receptor.　The　paper-based　Au+　disproportionation-mediated　AuNBP　morphology　system　not　only　has　excellent　stability,　reproducibility,　and　robust　resistibility　against　reducing　substances　existing　in　biological　samples　but　also　can　effectively　suppress　the　color　interference　from　reagents　and　greatly　improve　the　sensitivity　of　regulating　AuNBP　morphology　transformation,　making　it　capable　of　generating　long-term　stable　and　more　colorful/clearer　color　changes　corresponding　to　Au+　concentrations.　The　developed　PMS　exhibited　eight　color　changes　corresponding　to　AChE　activity　within　0.0-1.0　U/L　and　the　color　changes　could　be　stably　maintained　for　30　days,　which　makes　both　on-site　visual　detection　and　laboratory　sensitive　quantification　possible.　The　PMS　was　successfully　used　to　detect　AChE　activity　in　serum　and　erythrocyte　samples　with　a　visual　detection　limit　(LOD)　of　50　mU/L,　a　spectrometry　LOD　of　26　mU/L,　and　a　recovery　of　81-102%.　The　results　obtained　with　the　PMS　were　consistent　with　those　obtained　with　Ellman’s　method,　verifying　its　reliability.　All　of　the　above　features　make　the　PMS　a　promising　assay　for　the　rapid　detection　of　AChE　activity　in　clinical　diagnosis.　Importantly,　the　proposed　paper-based　Au+　disproportionation-mediated　AuNBP　morphology　system　can　serve　as　a　promising　general　signal　reporter　for　developing　various　sensitive　multicolor　colorimetric　sensors　for　target　detection　in　various　fields.　©　2024　American　Chemical　Society.