Electrogenerated chemiluminescence.: 80.: C-reactive protein determination at high amplification with [Ru(bpy)3]2+-containing microspheres

Biotinylated anti-C-reactive protein (CRP) species were attached to the surface of streptavidin-coated magnetic beads (MB) and avidin-coated polystyrene microspheres/ beads (PSB) entrapping a large number of electrogenerated chemiluminescence (ECL) labels (similar to10(9) Ru(bpy)(3)[B(C6F5)(4)](2)/bead) to form anti-CRP<---->MB and Ru(II)subset ofPSB/ avidin<---->anti-CRP conjugates, respectively. Sandwich-type RU(II)subset ofPSB/avidin<---->anti-CRP anti-CRP<---->MB aggregates were formed when Ru(II)subset ofPSB/avidin<---->anti-CRP was mixed with anti-CRP<---->MB conjugates in the presence of analyte CRP. The newly formed aggregates were magnetically separated from the reaction media and dissolved in MeCN containing tri-n-propylamine as an ECL coreactant. ECL was carried out with a potential scan from 0 to 2.8 V vs Ag/Ag+, and the ECL intensity was found to be proportional to the analyte CRP concentration over the range of 0.010-10 mug/mL. The CRP concentration of an unknown human plasma specimen was measured by the standard addition method based on this technique. Elimination of the nonspecific adsorption of the CRP system with several different blocking agents was also studied, and 2.0% bovine serum albumin was found to be best.