The　negative　charge　of　nanoparticles　affects　their　diffusion　toward　the　negatively　charged　indium　tin　oxide　(ITO)　electrode,　and　this　had　been　coupled　with　target-activated　CRISPR/Cas12a　to　develop　a　homogeneous　electrochemiluminescence　(ECL)　biosensor　for　human　papillomavirus　16　(HPV　16)　DNA　(chosen　as　model　target)　detection.　Tris(2,2′-bipyridyl)　ruthenium(II)　chloride　hexahydrate-doped　SiO2　nanoparticles　(Ru@SiO2　NPs)　with　a　diameter　of　about　50　nm　was　selected　as　the　ECL　probe.They　were　formed　by　encased　Ru(bpy)32+　in　SiO2　nanoparticles,　and　this　can　effectively　prevent　the　leakage　of　Ru(bpy)32+　molecules　and　improve　the　stability　of　the　probe.　A　hairpin　probe　(HP)　was　carefully　designed　and　modified　on　the　Ru@SiO2　NPs　surface　and　resulted　in　the　generation　of　a　large　negative　charge　on　the　surface　of　the　nanoparticles.　The　strong　electrostatic　repulsion　prevented　the　nanoparticles　from　reaching　the　negatively　charged　ITO　electrode,　leading　to　the　weakness　of　the　ECL　signal　detected.　In　the　presence　of　a　target,　Cas12a　is　activated　and　the　single-stranded　region　of　HP　is　cleaved　and　separated　from　the　Ru@SiO2　surface,　resulting　in　less　negative　charge　of　the　particles　and　enhanced　ECL　signal　of　the　system.　The　ECL　signal　was　linearly　correlated　with　the　logarithm　of　the　HPV　16　DNA　concentration　in　the　range　of　10　fM-10　nM　with　a　limit　of　3.4　fM　(S/N　=　3)　under　optimal　conditions.　The　biosensor　shows　exceptional　performance　in　the　detection　of　HPV16　DNA　in　real　samples.　©　2023　American　Chemical　Society.