Surface-adsorbed silver half-shells as a platform for surface-enhanced immunoassays; optimization through morphological control.

We have investigated the effectiveness of surface-adsorbed silver half-shells for inducing the surface-enhanced fluorescence phenomenon. Our simple structure consists of a dense monolayer of monodispersive latex sphere covered by thermally evaporated silver, some tens of nm thick. In order to increase its effectiveness as a platform for immunoassay, we explored three physical parameters; the diameter of the latex sphere, the deposition thickness and the adsorption density of the latex sphere. The maximum enhancement of 25.5 was achieved using the sphere with 56 nm diameter with 10 nm silver deposited. As for the adsorption density, the maximum fluorescence signal was achieved with a relatively sparse adsorption density, 12.5 per microm(2) vs. 105.9 per microm(2), corresponding to the full coverage. With the adsorption density also optimized, the enhancement was further increased by a factor of three.

Immunochemical detection of 3-deoxyglucosone in serum.

3-Deoxyglucosone (3-DG) is a metabolite of glucose that is thought to lead to the production of advanced glycation end products in diabetes. The previous assay for 3-DG in serum was based on a multi-step protocol, including derivatization, extraction, HPLC separation, and detection. In the current studies, we established a monoclonal antibody that recognizes the 3-DG-derivative, which is generated by the reaction of 3-DG and a 2,3-diamino-benzene derivative. Attachment of a biotin moiety to the 2,3-diamino-benzene ring via a linker allowed development of a highly sensitive chemiluminescent enzyme immunoassay for 3-DG equivalents. Unlike the previous assay, this method does not require extraction of 3-DG derivatives from serum. Treatment of 3-DG in serum with the DAB-link-biotin produced a quinoxaline derivative, which was specifically recognized by the monoclonal antibody. Using this assay, we found that serum 3-DG was higher in streptozotocin-induced diabetic rats than in normal control rats (25+/-5.6 vs. 9.8+/-1.1 microg/L). This simple assay may allow the monitoring of conditions leading to the accumulation of advanced glycation end products and evaluation of the risk of complications in diabetic patients.