title: Quantitative multianalyte microarray immunoassay utilizing upconverting phosphor technology.
abstract: A quantitative multianalyte immunoassay utilizing luminescent upconverting single-crystal nanoparticles as reporters on an antibody array-in-well platform was demonstrated. Upconverting nanoparticles are inorganic rare earth doped materials that have the unique feature of converting low energy infrared radiation into higher energy visible light. Autofluorescence, commonly limiting the sensitivity of fluorescence-based assays, can be completely eliminated with photon upconversion technology because the phenomenon does not occur in biological materials. Biotinylated antibodies for three analytes (prostate specific antigen, thyroid stimulating hormone, and luteinizing hormone) were printed in an array format onto the bottom of streptavidin-coated microtiter wells. Analyte dilutions were added to the wells, and the analytes were detected with antibody-coated upconverting nanoparticles. Binding of the upconverting nanoparticles was imaged with an anti-Stokes photoluminescence microwell imager, and the standard curves for each analyte were quantified from the selected spot areas of the images. Single analyte and reference assays were also carried out to compare with the results of the multianalyte assay. Multiplexing did not have an effect on the assay performance. This study demonstrates the feasibility of upconverting single-crystal nanoparticles for imaging-based detection of quantitative multianalyte assays.
authors.affiliation: Department of Biotechnology, University of Turku, Tykistökatu 6A, FI-20520 Turku, Finland. henna.pakkila@utu.fi
authors.forename: Henna
authors.initials: H
authors.lastname: Päkkilä
authors.forename: Minna
authors.initials: M
authors.lastname: Ylihärsilä
authors.forename: Satu
authors.initials: S
authors.lastname: Lahtinen
authors.forename: Liisa
authors.initials: L
authors.lastname: Hattara
authors.forename: Niina
authors.initials: N
authors.lastname: Salminen
authors.forename: Riikka
authors.initials: R
authors.lastname: Arppe
authors.forename: Mika
authors.initials: M
authors.lastname: Lastusaari
authors.forename: Petri
authors.initials: P
authors.lastname: Saviranta
authors.forename: Tero
authors.initials: T
authors.lastname: Soukka
mesh_terms: D000818:Animals; D055910:Antibodies, Immobilized / Q000737:chemistry / Q000276:immunology; D019921:Biotinylation; D005260:Female; D006801:Humans; D007118:Immunoassay / Q000295:instrumentation*; D049408:Luminescent Agents / Q000737:chemistry*; D008163:Luminescent Measurements / Q000295:instrumentation; D007986:Luteinizing Hormone / Q000032:analysis* / Q000276:immunology; D008297:Male; D053758:Nanoparticles / Q000737:chemistry*; D017430:Prostate-Specific Antigen / Q000032:analysis* / Q000276:immunology; D040081:Protein Array Analysis / Q000295:instrumentation; D012680:Sensitivity and Specificity; D013972:Thyrotropin / Q000032:analysis* / Q000276:immunology
chemical_list: D055910:Antibodies, Immobilized; D049408:Luminescent Agents; D007986:Luteinizing Hormone; D013972:Thyrotropin; D017430:Prostate-Specific Antigen