title: Refining the statistical model for quantitative immunostaining of surface-functionalized nanoparticles by AFM.
abstract: Recently, an atomic force microscopy (AFM)-based approach for quantifying the number of biological molecules conjugated to a nanoparticle surface at low number densities was reported. The number of target molecules conjugated to the analyte nanoparticle can be determined with single nanoparticle fidelity using antibody-mediated self-assembly to decorate the analyte nanoparticles with probe nanoparticles (i.e., quantitative immunostaining). This work refines the statistical models used to quantitatively interpret the observations when AFM is used to image the resulting structures. The refinements add terms to the previous statistical models to account for the physical sizes of the analyte nanoparticles, conjugated molecules, antibodies, and probe nanoparticles. Thus, a more physically realistic statistical computation can be implemented for a given sample of known qualitative composition, using the software scripts provided. Example AFM data sets, using horseradish peroxidase conjugated to gold nanoparticles, are presented to illustrate how to implement this method successfully.
authors.affiliation: Materials Measurements Science Division, Material Measurement Laboratory, National Institute of Standards and Technology, Gaithersburg, MD, 20899, USA, robert.maccuspie@nist.gov.
authors.forename: Robert I
authors.initials: RI
authors.lastname: MacCuspie
authors.forename: Danielle E
authors.initials: DE
authors.lastname: Gorka
mesh_terms: D031215:Armoracia / Q000201:enzymology; D004800:Enzymes, Immobilized / Q000032:analysis; D006046:Gold / Q000032:analysis; D006735:Horseradish Peroxidase / Q000032:analysis; D007074:Immunoglobulin G / Q000032:analysis; D007150:Immunohistochemistry / Q000379:methods*; D018625:Microscopy, Atomic Force / Q000379:methods*; D015233:Models, Statistical; D053758:Nanoparticles / Q000032:analysis* / Q000648:ultrastructure; D013499:Surface Properties
chemical_list: D004800:Enzymes, Immobilized; D007074:Immunoglobulin G; D006046:Gold; D006735:Horseradish Peroxidase