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The pig pancreas is considered to be the most suitable source of islets for xenotransplantation in patients with type I diabetes. The objective of this study was to assess the antigenicity of neonatal porcine islet-like cell clusters (NPCC), including the Galα1–3Galβ1–4GlcNAc-R (α-Gal) and Hanganutziu–Deicher (H–D) antigens, and the pathway involved in human complement activation. The efficiency of expression of human decay-accelerating factor (DAF: CD55) on NPCC by adenoviral transduction was also examined, and the functional capacity of DAF was also estimated.The deposition of human natural antibodies, immunoglobulin (Ig)G and IgM, and the expression of α-Gal and H–D antigens on NPCC were investigated by FACS analysis. The downregulation in the antigenicity to human natural antibodies, including the α-Gal and H–D antigens on NPCC by treatment with tunicamycin, PDMP and neuraminidase were also examined. In addition, complement-mediated islet lysis was examined using factor D-deficient and C1-deficient sera. An adenovirus encoding DAF under the control of the cytomegalovirus promoter, Ad.pCMV-DAF, was then constructed, and used for transducing NPCC. The amelioration of complement-dependent cytotoxicity of the NPCC by the transduced DAF was assessed as an in vitro hyperacute rejection model of a pig to human xenograft.The NPCC clearly expressed the α-Gal epitope, and the human natural antibodies, IgG and IgM, and the anti-H–D antibody also reacted with the NPCC. Treatment of NPCC with tunicamycin led to a drastic reduction in the extent of deposition of IgG, indicating the importance of N-linked sugars on the islets, presumably related to α-Gal expression on N-linked sugars. Neuraminidase treatment indicated the presence of, not only the H–D antigen, but also other sialic acid antigens which reacted with the human natural antibody, especially IgG. The complement deposition of factor B on NPCC was clear, and the alternative pathway-mediated NPCC killing accounted for approximately 30% of that by the total complement pathway. On the other hand, approximately 90% of the NPCC could be transduced to express DAF by the adenovector, Ad.pCMV-DAF. The expressed DAF showed an approximately 50–62% suppression in complement-dependent NPCC lysis.The origin of the antigenicity of NPCC is mainly N-linked sugars including α-Gal and sialic acid antigens, and NPCC expressed the transduced molecule in high efficiency by the adenovector.