Human　serum　albumin　(HSA),　a　naturally　abundant　protein　in　blood　plasma　and　tissue　fluids,　has　an　extraordinary　ligand-binding　capacity　and　is　advocated　as　a　drug　carrier　to　facilitate　drug　delivery.　To　render　it　tumor　targeting　specificity,　we　generated　a　recombinant　HSA　fused　with　the　amino-　terminal　fragment　(ATF)　of　urokinase,　allowing　the　fusion　protein　to　bind　to　urokinase　receptor　(uPAR),　which　is　shown　to　have　a　high　expression　level　in　many　tumors,　but　not　in　normal　tissues.　To　test　the　efficacy　of　this　bifunctional　protein　(ATF-HSA),　a　hydrophobic　photosensitizer　(mono-substituted　β-carboxy　phthalocyanine　zinc,　CPZ)　was　chosen　as　a　cytotoxic　agent.　A　dilution-　incubation-purification　(DIP)　strategy　was　developed　to　load　the　ATF-HSA　with　this　CPZ,　forming　a　1:1　molecular　complex　(ATF-HSA:CPZ).　We　demonstrated　that　CPZ　was　indeed　embedded　inside　ATF-HSA　at　the　fatty　acid　binding　site　1　(FA1)　of　HSA,　giving　a　hydrodynamic　radius　of　7.5　nm,　close　to　HSA＇s　(6.5　nm).　ATF-HSA:CPZ　showed　high　stability　and　remarkable　optical　and　photophysical　properties　in　aqueous　solution.　In　addition,　the　molecular　complex　ATF-HSA:CPZ　can　bind　to　recombinant　uPAR　in　vitro　and　uPAR　on　tumor　cell　surfaces,　and　was　efficient　in　photodynamic　killing　of　tumor　cells.　The　tumor-killing　potency　of　this　molecular　complex　was　further　demonstrated　in　a　tumor-bearing　mouse　model　at　a　dose　of　0.080　μmol　/　kg,　or　0.050　mg　CPZ　/　kg　of　mouse　body　weight.　Using　fluorescent　molecular　tomography　(FMT),　ATF-HSA:CPZ　was　shown　to　accumulate　specifically　in　tumors,　and　importantly,　such　tumor　retention　was　higher　than　that　of　HSA:CPZ.　Together,　these　results　indicate　that　ATF-HSA:CPZ　is　not　only　an　efficient　tumor-specific　cytotoxic　agent,　but　also　an　useful　tumor-specific　imaging　probe.　This　bifunctional　protein　ATF-HSA　can　also　be　used　as　a　drug　carrier　for　other　types　of　cytotoxic　or　imaging　agents　to　render　them　specificity　for　uPAR-expressing　tumors.　©　Ivyspring　International　Publisher.