Quantifying the Numbers of Gold Nanoparticles in the Test Zone of Lateral-Flow Immunoassay Strips

Lateral flow immunoassay (LFIA) is a rapid and simple point-of-care method for the detection of various analytes. In the colorimetric sandwich format, the reaction outcome is a colored test zone line formed by Au nanoparticle (AuNP) conjugates captured by bound analyte molecules. Although nanoparticle design is crucial for the assay sensitivity, the correlation between the test zone brightness and the number and size of captured AuNPs has not been studied in detail. To fill this gap, we used an unprecedented set of 10 spherical and monodisperse AuNPs with diameters d ranging from 16 to 115 nm. The calculated optical properties are in excellent agreement with two-layered Mie theory for Au cores coated with 3-nm CTAC shells with a refractive index of 1.5. Different concentrations of AuNPs (ICP-MS and UV-vis measurements) were spotted onto a nitrocellulose LFIA membrane, and the color intensity of the spots was measured and analyzed with RGB and HSV color parameters. The minimal detected spot intensity was p...