A　novel　fluoresencent　immunosensor　for　determination　of　cancer　biomarkers　such　as　alpha-fetoprotein　(AFP)　was　designed　by　utilizing　both　the　high　specificity　of　antigen-antibody　sandwich　structure　and　the　high　sensitivity　of　the　click　chemistry　based　fluorescence　detection.　Instead　of　an　enzyme　or　fluorophore,　the　CuO　nanoparticles　are　labeled　on　the　detection　antibody,　which　was　not　susceptible　to　the　change　of　the　external　environments.　The　CuO　nanoparticles　which　were　modified　on　the　sandwich　structure　can　be　dissolved　to　produce　Cu2+　ions　with　the　help　of　HCl　and　then　the　Cu2+　ions　were　reduced　by　sodium　ascorbate　to　produce　Cu+　ions　which　triggered　the　Cu+　catalyzed　alkyne-azide　cycloaddition　(CuAAC)　reaction　between　the　weak　fluorescent　compound　(3-azido-7-hydroxycoumarin)　and　propargyl　alcohol　to　form　a　strong　fluorescent　compound.　A　good　linear　relationship　was　observed　between　the　fluorescence　increase　factor　of　the　system　and　the　concentration　of　AFP　in　the　range　of　0.025-5.0　ng/mL　with　a　detection　limit　of　12　pg/mL　(S/N=3).　The　proposed　fluorescent　sensor　had　been　applied　to　detect　AFP　in　the　human　serum　samples　and　gave　satisfactory　results.　(C)　2015　Elsevier　B.V.　All　rights　reserved.