title: Sensitive targeted multiple protein quantification based on elemental detection of quantum dots.
abstract: A generic strategy based on the use of CdSe/ZnS Quantum Dots (QDs) as elemental labels for protein quantification, using immunoassays with elemental mass spectrometry (ICP-MS), detection is presented. In this strategy, streptavidin modified QDs (QDs-SA) are bioconjugated to a biotinylated secondary antibody (b-Ab2). After a multi-technique characterization of the synthesized generic platform (QDs-SA-b-Ab2) it was applied to the sequential quantification of five proteins (transferrin, complement C3, apolipoprotein A1, transthyretin and apolipoprotein A4) at different concentration levels in human serum samples. It is shown how this generic strategy does only require the appropriate unlabeled primary antibody for each protein to be detected. Therefore, it introduces a way out to the need for the cumbersome and specific bioconjugation of the QDs to the corresponding specific recognition antibody for every target analyte (protein). Results obtained were validated with those obtained using UV-vis spectrophotometry and commercial ELISA Kits. As expected, ICP-MS offered one order of magnitude lower DL (0.23 fmol absolute for transferrin) than the classical spectrophotometric detection (3.2 fmol absolute). ICP-MS precision and detection limits, however turned out to be compromised by procedural blanks. The full analytical performance of the ICP-MS-based immunoassay proposed was assessed for detection of transferrin (Tf), present at the low ng mL(-1) range in a complex "model" synthetic matrix, where the total protein concentration was 100 μg mL(-1). Finally, ICP-MS detection allowed the quantitative control of all the steps of the proposed immunoassay, by computing mass balances obtained, and the development of a faster indirect immunoassay format where the plate wells were directly coated with the whole protein mixture sample.
authors.affiliation: Department of Physical and Analytical Chemistry, University of Oviedo, Julián Clavería 8, Oviedo 33006, Spain.
authors.forename: Antonio R
authors.initials: AR
authors.lastname: Montoro Bustos
authors.affiliation: Department of Physical and Analytical Chemistry, University of Oviedo, Julián Clavería 8, Oviedo 33006, Spain.
authors.forename: Marta
authors.initials: M
authors.lastname: Garcia-Cortes
authors.affiliation: Fundación de Investigación Oftalmológica, Instituto Oftalmológico Fernandez-Vega, Avenida Doctores Fernández-Vega, 34, Oviedo 33012, Spain.
authors.forename: Hector
authors.initials: H
authors.lastname: González-Iglesias
authors.affiliation: Department of Physical and Analytical Chemistry, University of Oviedo, Julián Clavería 8, Oviedo 33006, Spain. Electronic address: ruizjorge@uniovi.es.
authors.forename: Jorge
authors.initials: J
authors.lastname: Ruiz Encinar
authors.affiliation: Department of Physical and Analytical Chemistry, University of Oviedo, Julián Clavería 8, Oviedo 33006, Spain.
authors.forename: José M
authors.initials: JM
authors.lastname: Costa-Fernández
authors.affiliation: Fundación de Investigación Oftalmológica, Instituto Oftalmológico Fernandez-Vega, Avenida Doctores Fernández-Vega, 34, Oviedo 33012, Spain; Department of Ophthalmology and Visual Science, Yale University School of Medicine, New Haven, CT 06510, USA.
authors.forename: Miguel
authors.initials: M
authors.lastname: Coca-Prados
authors.affiliation: Department of Physical and Analytical Chemistry, University of Oviedo, Julián Clavería 8, Oviedo 33006, Spain. Electronic address: asm@uniovi.es.
authors.forename: Alfredo
authors.initials: A
authors.lastname: Sanz-Medel
mesh_terms: D001798:Blood Proteins / Q000032:analysis*; D019187:Cadmium Compounds / Q000737:chemistry; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D057230:Limit of Detection; D013058:Mass Spectrometry / Q000379:methods*; D045663:Quantum Dots / Q000737:chemistry*; D015203:Reproducibility of Results; D018036:Selenium Compounds / Q000737:chemistry; D013440:Sulfides / Q000737:chemistry; D017967:Zinc Compounds / Q000737:chemistry
chemical_list: D001798:Blood Proteins; D019187:Cadmium Compounds; D018036:Selenium Compounds; D013440:Sulfides; D017967:Zinc Compounds; C058667:cadmium selenide; C031238:zinc sulfide
keywords: Elemental mass spectrometry; Immunoassay; Labeling; Protein quantification; Quantum Dots