Ultrasensitive amperometric magnetoimmunosensor for human C-reactive protein quantification in serum

A highly sensitive magnetoimmunosensor for the determination of human C-reactive protein (CRP) is described in this work. A sandwich format involving covalent immobilization of the capture antibody (antiCRP) onto carboxylic-modified magnetic beads (HOOC-MBs) activated with N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide (EDC) and N-hydroxysulfosuccinimide (sulfo-NHS), the antigen-antibody reaction and incubation of the modified MBs with a biotynilated antibody (biotin-antiCRP), is used. Furthermore, an incubation step with a Streptavidin HRP (Strp-HRP) conjugate is employed to allow monitoring of the affinity reaction. The resulting modified MBs are captured by a magnet placed under the surface of a disposable gold screen-printed electrode (Au/SPE) and the amperometric responses are measured at −0.10 V (vs. a Ag pseudo-reference electrode), upon addition of 3,3′,5,5′-tetramethylbenzidine (TMB) as electron transfer mediator and H2O2 as the enzyme substrate. The CRP magnetoimmunosensor exhibited a wide range of linearity between 0.07 and 1000 ng mL−1 with a low detection limit of (0.021 ± 0.005) ng mL−1. The magnetoimmunosensor was successfully applied to the analysis of an international standard for CRP serum.