Electrochemical Immunoassay for Mucin 1 Detection as a Diagnostic Tool in Ovarian Cancer
The immunoassay developed for Mucin 1 (MUC1) detection is based on a sandwich format in which a primary antibody immobilized on the surface of magnetic beads specifically binds the MUC1 protein. The sandwich immunoassay is performed by adding secondary an
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Introduction
Cancer is one of the leading causes of death worldwide. Among this, ovarian cancer represents the fifth cause of death and the most lethal of all gynecological malignancies [1, 2]. Early detection remains the most promising approach to improve longterm survival rate of patients affected by cancer [3]. For this, the analysis of the cancer biomarker represents an ideal tool to determine diagnosis and prognosis and follows the medical treatment of patients [4–6]. MUC1 gene encodes a type I transmembrane glycoprotein that is expressed on the apical surface of various epithelial cells. In malignant neoplasm, Mucin 1 (MUC1) loses its apical distribution, becomes underglycosylated and overexpressed, and is secreted into the blood circulation, serving therefore as potential tumor marker and prognosis factor in different types of cancer [7]. In this paper an electrochemical immunoassay as a screening device for the detection of MUC1 in real samples was presented. The proposed approach uses disposable screen-printed electrodes as transducers and a simple target capturing step by antibody-functionalized magnetic beads.
A. Ravalli • G. Marrazza (*) Department of Chemistry “Ugo Schiff”, University of Florence, Via della Lastruccia 3, 50019 Sesto Fiorentino, Florence, Italy e-mail: [email protected] A. Florea • C. Cristea • R. Sandulescu Department of Analytical Chemistry, Faculty of Pharmacy, University of Medicine and Pharmacy, “Iuliu Hatieganu”, Pasteur 4, Cluj-Napoca, Romania C. Di Natale et al. (eds.), Sensors and Microsystems: Proceedings of the 17th National Conference, Brescia, Italy, 5-7 February 2013, Lecture Notes in Electrical Engineering 268, DOI 10.1007/978-3-319-00684-0_31, © Springer International Publishing Switzerland 2014
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Materials and Methods
Dynabeads Protein G-coated magnetic beads were purchased from Invitrogen (Milan, Italy). MUC1 protein, MUC1 monoclonal mouse antibody (Ab1), MUC1 polyclonal rabbit antibody (Ab2), and polyclonal antibody anti-rabbit IgG labeled with alkaline phosphatase (Ab3-AP) were provided by Novus Biological (Cambridge, UK). 1-Naphthyl phosphate, diethanolamine, potassium chloride, magnesium chloride, and commercial non-pathological human serum were purchased from SigmaAldrich (Milan, Italy). All solutions were prepared using water from Milli-Q Water Purification System (Millipore, UK). Eight screen-printed cells were used in the experiments. Each cell is based on graphite working electrode (2.0 mm in diameter), each with a graphite counter electrode and a silver pseudo-reference electrode, produced on a DEK 248 (DEK, Weymouth, UK) screen printing machine [8]. Electrochemical measurements were performed with μAutolab type II PGSTAT (Metrohm, The Netherlands) with General Purpose Electrochemical System (GPES) 4.9 software and PalmSens handheld potentiostat (PalmSens BV, The Netherlands). Differential pulse voltammetry (DPV) was employed as electrochemical technique using the following parameters: potential range of −0.2 to
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