title: Sensitivity enhancement of surface plasmon resonance biosensing of small molecules.
abstract: Surface plasmon resonance (SPR) biosensor formats using gold nanoparticle or protein signal amplification for the sensitive assay of small molecules were developed using progesterone as a model compound. Progesterone was immobilized to a dextran surface in the Biacore biosensor through in situ covalent immobilization using an oligoethylene glycol linker attached to the 4 position of the steroid. This surface produced stable antibody binding for in excess of 1100 assay cycles. Using this surface, assays were developed for progesterone using 10- and 20-nm gold-streptavidin labels attached to biotinylated monoclonal antibody in both label prebinding and sequential binding formats. Prelabeling formats gave no signal enhancement but produced assays with limits of detection of 143 pg/ml, compared with approximately 1 ng/ml in previous studies. Sequential binding formats gave signal enhancements of 2.2-fold over the monoclonal antibody and a limit of detection of 23.1 pg/ml. It was found that secondary antibody labeling gave 8.1-fold signal enhancements and a limit of detection of 20.1 pg/ml, whereas use of secondary antibody-25 nm gold complexes provided more signal enhancement (13-fold) and a further improvement in limit of detection of 8.6 pg/ml.
authors.affiliation: Bioengineering Sector, HortResearch, Private Bag 3123, Hamilton, New Zealand.
authors.forename: John S
authors.initials: JS
authors.lastname: Mitchell
authors.forename: Yinqiu
authors.initials: Y
authors.lastname: Wu
authors.forename: Christian J
authors.initials: CJ
authors.lastname: Cook
authors.forename: Lyndsay
authors.initials: L
authors.lastname: Main
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry; D017979:Gold Colloid / Q000737:chemistry; D011374:Progesterone / Q000032:analysis*; D012680:Sensitivity and Specificity; D020349:Surface Plasmon Resonance / Q000379:methods
chemical_list: D000911:Antibodies, Monoclonal; D017979:Gold Colloid; D011374:Progesterone