In　this　study,　an　electrochemiluminescence　resonance　energy　transfer　(ECL-RET)　biosensor　with　high　sensitivity　and　strong　resistance　to　interference　was　constructed　based　on　the　CRISPR/Cas13a　system　and　magnetic　separation　for　ovarian　cancer　biomarker　miR-543　detection.　Mesoporous　silica　nanoparticles　embedded　with　Ru(bpy)32+　(Ru@SiO2)　have　high　electrochemiluminescence　(ECL)　response　was　chosen　as　energy　donor.　Single-stranded　DNA　S1　containing　“rUrU”　motif　was　immobilized　on　AuNRs　(AuNRs-S1),　which　hybridized　with　single-stranded　DNA　S2　modified　SAMBs　(SAMBs-S2)　to　form　AuNRs-S1/S2-SAMBs　complex,　this　has　been　used　as　energy　acceptor.　In　the　absence　of　the　target,　Cas13a　remained　inactive,　preventing　the　cleavage　of　S1,　thereby　maintaining　the　association　of　AuNRs　with　SAMBs.　Then　they　were　added　in　Ru@SiO2　solution　after　magnetic　separation.　The　electrostatic　adsorption　between　the　negatively　charged　AuNRs　and　the　positively　charged　Ru@SiO2　cause　the　occurrence　of　ECL-RET　and　low　ECL　signal　had　been　detected.　When　the　target　was　added,　Cas13a　was　activated　and　resulted　in　the　non-specifically　cleaving　of　S1,　so　AuNRs　detached　from　SAMBs.　After　magnetic　separation,　fewer　AuNRs　participated　in　ECL-RET,　leading　to　an　enhanced　ECL　signal　detected.　The　change　in　ECL　intensity　(ΔECL)　exhibited　a　linear　correlation　with　the　logarithm　of　miR-543　concentration　within　the　range　of　10　fM　to　10　nM,　with　a　detection　limit　of　6.91　fM.　The　biosensor　had　been　applied　to　detect　miR-543　in　clinical　samples　with　high　accuracy.　©　2025　Elsevier　B.V.