Amperometric immunoassay for the obesity biomarker amylin using a screen printed carbon electrode functionalized with an electropolymerized carboxylated polypyrrole
- 227 Downloads
Amylin (the islet amyloid polypeptide) is a hormone related to adiposity, hunger and satiety. It is co-secreted with insulin from pancreatic B-cells. An amperometric immunosensor is presented here for the determination of amylin. It is making use of a screen printed carbon electrode (SPCE) functionalized with electropolymerized poly(pyrrole propionic acid) (pPPA) with abundant carboxyl groups that facilitate covalent binding of antibody against amylin. A competitive immunoassay was implemented using biotinylated amylin and streptavidin labeled with horse radish peroxidase (HRP-Strept) as the enzymatic tracer. The amperometric detection of H2O2 mediated by hydroquinone was employed as an electrochemical probe to monitor the affinity reaction. The variables involved in the preparation and function of the immunosensor were optimized and the electrodes were characterized by electrochemical impedance spectroscopy and cyclic voltammetry. The calibration graph for amylin, obtained by amperometry at −200 mV vs Ag pseudo-reference electrode, showed a range of linearity extending from 1.0 fg∙mL−1 to 50 pg∙mL−1, with a detection limit of 0.92 fg∙mL−1. This is approximately 7000 times lower than the minimum detectable concentration reported for the ELISA immunoassays available for amylin. The assay has excellent reproducibility and good selectivity over potential interferents.
KeywordsScreen-printed carbon electrodes Electrochemical biosensor Conducting polymer Mix&Go™ Urine Serum
The financial support of projects CTQ2015-70023-R (Spanish Ministry of Economy and Competitivity Research Projects), and S2013/MT-3029 (NANOAVANSENS Program from the Comunidad de Madrid) are gratefully acknowledged.
Compliance with ethical standards
The authors declare that they have no competing interest.
- 7.Sanke T, Hanabusa T, Nakano Y, Oki C, Okai K, Nishimura S, Kondo M, Nanjo K (1991) Plasma islet amyloid polypeptide (amylin) levels and their responses to oral glucose in type-2 (non-insulin-dependent) diabetic patients. Diabetologia 34:129–132. https://doi.org/10.1007/BF00500385 CrossRefPubMedGoogle Scholar
- 12.Ojeda I, Barrejón M, Arellano LM, González-Cortés A, Yáñez-Sedeño P, Langa F, Pingarrón JM (2015) Grafted-double walled carbon nanotubes as electrochemical platforms for immobilization of antibodies using a metallic-complex chelating polymer: application to the determination of adiponectin cytokine in serum. Biosens Bioelectron 74:24–29. https://doi.org/10.1016/j.bios.2015.06.001 CrossRefPubMedGoogle Scholar
- 14.Serafín V, Torrente-Rodríguez RM, Batlle M, García de Frutos P, Campuzano S, Yáñez-Sedeño P, Pingarrón JM (2017) Electrochemical immunosensor for receptor tyrosine kinase AXL using poly(pyrrolepropionic acid)-modified disposable electrodes. Sensors Actuators B Chem 240:1251–1256. https://doi.org/10.1016/j.snb.2016.09.109 CrossRefGoogle Scholar
- 15.Eguílaz M, Moreno-Guzmán M, Campuzano S, González-Cortés A, Yáñez-Sedeño P, Pingarrón JM (2010) An electrochemical immunosensor for testosterone using functionalized magnetic beads and screen-printed carbon electrodes. Biosens Bioelectron 26:517–522. https://doi.org/10.1016/j.bios.2010.07.060 CrossRefPubMedGoogle Scholar
- 16.Dong H, Cao X, Li CM, Hu W (2008) An in situ electrochemical surface plasmon resonance immunosensor with polypyrrole propylic acid film: comparison between SPR and electrochemical responses from polymer formation to protein immunosensing. Biosens Bioelectron 23:1055–1062. https://doi.org/10.1016/j.bios.2007.10.026 CrossRefPubMedGoogle Scholar
- 17.Ball GM, in "Vitamins in foods: analysis, Bioavalaibility, and stability", CRC Taylor and Francis, Boca Raton FL (2006) p. 223.Google Scholar