title: The development of quantum dot calibration beads and quantitative multicolor bioassays in flow cytometry and microscopy.
abstract: The use of fluorescence calibration beads has been the hallmark of quantitative flow cytometry. It has enabled the direct comparison of interlaboratory data as well as quality control in clinical flow cytometry. In this article, we describe a simple method for producing color-generalizable calibration beads based on streptavidin functionalized quantum dots. Based on their broad absorption spectra and relatively narrow emission, which is tunable on the basis of dot size, quantum dot calibration beads can be made for any fluorophore that matches their emission color. In an earlier publication, we characterized the spectroscopic properties of commercial streptavidin functionalized dots (Invitrogen). Here we describe the molecular assembly of these dots on biotinylated beads. The law of mass action is used to readily define the site densities of the dots on the beads. The applicability of these beads is tested against the industry standard, namely commercial fluorescein calibration beads. The utility of the calibration beads is also extended to the characterization surface densities of dot-labeled epidermal growth factor ligands as well as quantitative indicators of the binding of dot-labeled virus particles to cells.
authors.affiliation: Department of Pathology and Cancer Center, University of New Mexico, Albuquerque, NM 87131, USA.
authors.forename: Yang
authors.initials: Y
authors.lastname: Wu
authors.forename: Samuel K
authors.initials: SK
authors.lastname: Campos
authors.forename: Gabriel P
authors.initials: GP
authors.lastname: Lopez
authors.forename: Michelle A
authors.initials: MA
authors.lastname: Ozbun
authors.forename: Larry A
authors.initials: LA
authors.lastname: Sklar
authors.forename: Tione
authors.initials: T
authors.lastname: Buranda
mesh_terms: D015374:Biosensing Techniques / Q000379:methods*; D001710:Biotin / Q000737:chemistry; D002138:Calibration / Q000592:standards; D005069:Evaluation Studies as Topic; D005434:Flow Cytometry / Q000379:methods / Q000592:standards*; D005455:Fluorescent Antibody Technique / Q000592:standards; D005456:Fluorescent Dyes / Q000737:chemistry / Q000592:standards; D006801:Humans; D008856:Microscopy, Fluorescence / Q000379:methods / Q000592:standards*; D008863:Microspheres; D015336:Molecular Probe Techniques; D036103:Nanotechnology / Q000379:methods*; D045663:Quantum Dots; D012015:Reference Standards; D019809:Streptavidin / Q000737:chemistry; D014407:Tumor Cells, Cultured
chemical_list: D005456:Fluorescent Dyes; D001710:Biotin; D019809:Streptavidin