title: A novel sensitive immunoassay method based on the Invader technique.
abstract: A novel and sensitive immunoassay method has been developed in which the conventional sandwich immunoassay and the highly sensitive DNA detection method, the Invader method, are combined. The signal amplification function of the latter method has been successfully used to enhance the sensitivity of the sandwich immunoassay. The new assay method may be called the Immuno-Invader assay. The assay format involves three important steps: (1) a target antigen is captured and flagged with a biotin-conjugated detection antibody by the sandwich method, (2) streptavidin and a biotin-conjugated oligonucleotide are added to form a complex with the detection antibody, and (3) the oligonucleotide in the complex is detected using the Invader method. The method was applied to the assay of human tumor necrosis factor-alpha (hTNF-alpha). Detection limits obtained were 0.1 pg/ml hTNF-alpha when a luminescent europium chelate was used with a time-resolved measurement mode, and 0.8 pg/ml when fluorescein was used with a normal prompt fluorescence measurement mode. On the other hand, the detection limit of a commercially available hTNF-alpha enzyme-linked immunosorbent assay that uses horseradish peroxidase was 3.5 pg/ml. These results demonstrate the feasibility and potential of the new assay method for highly sensitive immunoassay.
authors.affiliation: Pre-venture Program, Office of Technology Transfer, Japan Science and Technology Agency, 5-3 Yonbancho, Chiyoda-ku, Tokyo 102-8666, Japan.
authors.forename: Min-Jue
authors.initials: MJ
authors.lastname: Xie
authors.forename: Kôichi
authors.initials: K
authors.lastname: Fukui
authors.forename: Masanori
authors.initials: M
authors.lastname: Horie
authors.forename: Yuri
authors.initials: Y
authors.lastname: Sakihama
authors.forename: Kimikazu
authors.initials: K
authors.lastname: Hashino
authors.forename: Hiroko
authors.initials: H
authors.lastname: Kimura
authors.forename: Kazuko
authors.initials: K
authors.lastname: Matsumoto
mesh_terms: D004797:Enzyme-Linked Immunosorbent Assay; D005063:Europium / Q000737:chemistry; D019793:Fluorescein / Q000737:chemistry; D005453:Fluorescence; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes / Q000737:chemistry; D006735:Horseradish Peroxidase / Q000032:analysis / Q000276:immunology; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D012680:Sensitivity and Specificity; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis / Q000276:immunology
chemical_list: D005456:Fluorescent Dyes; D014409:Tumor Necrosis Factor-alpha; D005063:Europium; D006735:Horseradish Peroxidase; D019793:Fluorescein