title: Super-sensitive time-resolved fluoroimmunoassay for thyroid-stimulating hormone utilizing europium(III) nanoparticle labels achieved by protein corona stabilization, short binding time, and serum preprocessing.
abstract: Thyrotropin or thyroid-stimulating hormone (TSH) is used as a marker for thyroid function. More precise and more sensitive immunoassays are needed to facilitate continuous monitoring of thyroid dysfunctions and to assess the efficacy of the selected therapy and dosage of medication. Moreover, most thyroid diseases are autoimmune diseases making TSH assays very prone to immunoassay interferences due to autoantibodies in the sample matrix. We have developed a super-sensitive TSH immunoassay utilizing nanoparticle labels with a detection limit of 60 nU L-1in preprocessed serum samples by reducing nonspecific binding. The developed preprocessing step by affinity purification removed interfering compounds and improved the recovery of spiked TSH from serum. The sensitivity enhancement was achieved by stabilization of the protein corona of the nanoparticle bioconjugates and a spot-coated configuration of the active solid-phase that reduced sedimentation of the nanoparticle bioconjugates and their contact time with antibody-coated solid phase, thus making use of the higher association rate of specific binding due to high avidity nanoparticle bioconjugates. Graphical Abstract We were able to decrease the lowest limit of detection and increase sensitivity of TSH immunoassay using Eu(III)-nanoparticles. The improvement was achieved by decreasing binding time of nanoparticle bioconjugates by small capture area and fast circular rotation. Also, we applied a step to stabilize protein corona of the nanoparticles and a serum-preprocessing step with a structurally related antibody.
authors.affiliation: Laboratory of Biophysics, Institute of Biomedicine and Medicity research laboratories, University of Turku, Tykistökatu 6A, 20520, Turku, Finland. tuomas.nareoja@ki.se.|Division of Pathology, Department of Laboratory Medicine, Karolinska Institutet, F46, Karolinska Universitetssjukhuset, Huddinge, 141 86, Stockholm, Sweden. tuomas.nareoja@ki.se.
authors.forename: Tuomas
authors.initials: T
authors.lastname: Näreoja
authors.affiliation: Pharmaceutical Sciences Laboratory, Faculty of science and engineering, Åbo akademi University, Tykistökatu 6A, 20520, Turku, Finland.
authors.forename: Jessica M
authors.initials: JM
authors.lastname: Rosenholm
authors.affiliation: Department of Biochemistry/Biotechnology, University of Turku, Vatselankatu 2, 20500, Turku, Finland.
authors.forename: Urpo
authors.initials: U
authors.lastname: Lamminmäki
authors.affiliation: Laboratory of Biophysics, Institute of Biomedicine and Medicity research laboratories, University of Turku, Tykistökatu 6A, 20520, Turku, Finland.
authors.forename: Pekka E
authors.initials: PE
authors.lastname: Hänninen
mesh_terms: D015415:Biomarkers; D005063:Europium / Q000737:chemistry*; D015200:Fluoroimmunoassay / Q000379:methods*; D006801:Humans; D053768:Metal Nanoparticles / Q000737:chemistry*; D000066970:Protein Corona; D012680:Sensitivity and Specificity; D013194:Staining and Labeling; D013972:Thyrotropin / Q000097:blood*; D013997:Time Factors
chemical_list: D015415:Biomarkers; D000066970:Protein Corona; D005063:Europium; D013972:Thyrotropin
keywords: Immunoassay interference; Nanoparticle bioconjugate; Nanoparticle protein corona; Sandwich-type immunoassay; Time-resolved fluoroimmunoassay