Detection of the Inflammation Biomarker C-Reactive Protein in Serum Samples: Towards an Optimal Biosensor Formula

Author： Fakanya Wellington M. Tothill Ibtisam E.

Publisher： MDPI

E-ISSN： 2079-6374|4|4|340-357

ISSN： 2079-6374

Source： Biosensors, Vol.4, Iss.4, 2014-10, pp. : 340-357

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Abstract

The development of an electrochemical immunosensor for the biomarker, C-reactive protein (CRP), is reported in this work. CRP has been used to assess inflammation and is also used in a multi-biomarker system as a predictive biomarker for cardiovascular disease risk. A gold-based working electrode sensor was developed, and the types of electrode printing inks and ink curing techniques were then optimized. The electrodes with the best performance parameters were then employed for the construction of an immunosensor for CRP by immobilizing anti-human CRP antibody on the working electrode surface. A sandwich enzyme-linked immunosorbent assay (ELISA) was then constructed after sample addition by using anti-human CRP antibody labelled with horseradish peroxidase (HRP). The signal was generated by the addition of a mediator/substrate system comprised of 3,3,5',5'-Tetramethylbenzidine dihydrochloride (TMB) and hydrogen peroxide (H₂O₂). Measurements were conducted using chronoamperometry at −200 mV against an integrated Ag/AgCl reference electrode. A CRP limit of detection (LOD) of 2.2 ng·mL⁻¹ was achieved in spiked serum samples, and performance agreement was obtained with reference to a commercial ELISA kit. The developed CRP immunosensor was able to detect a diagnostically relevant range of the biomarker in serum without the need for signal amplification using nanoparticles, paving the way for future development on a cardiac panel electrochemical point-of-care diagnostic device.