Abstract
In this paper, an electrochemical magneto-immunosensor for the detection of human growth hormone (hGH) is described for the first time. The immunosensor involves the use of tosyl-activated magnetic microparticles (TsMBs) to covalently immobilize a monoclonal mAbhHG antibody. A sandwich-type immunoassay with a secondary pAbhGH antibody and anti-IgG labelled with alkaline phosphatase (anti-IgG-AP) was employed. TsMBs–mAbhGH–hGH–pAbhGH–anti-IgG-AP conjugates were deposited onto the surface of a screen-printed gold electrode using a small neodymium magnet, and electrochemical detection was performed by square-wave voltammetry upon the addition of 4-aminophenyl phosphate as the AP substrate. All the variables involved in the preparation of immunoconjugates and in the immunoassay protocol were optimized. A calibration curve for hGH was constructed with a linear range between 0.01 and 100 ng/mL (r = 0.998) and a limit of detection of 0.005 ng/mL. This value is nearly three orders of magnitude lower than that obtained using surface plasmon resonance (Treviño et al., Talanta 78:1011–1016, 2009). Furthermore, good repeatability, with RSD = 3% (n = 10) at the 1-ng/mL hGH level, was obtained. Cross-reactivity studies with other hormones demonstrated good selectivity. The magneto-immunosensor was applied to the analysis of human serum spiked with hGH at the 4- and 0.1-ng/mL levels. Mean recoveries of 96 ± 6% and 99 ± 2%, respectively, were obtained.
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Financial support from the Ministerio de Ciencia e Innovación (Projects DPS2008-07005-C02-01 and CTQ2009-12650), as well as of Comunidad de Madrid S2009/PPQ-1642, programme AVANSENS, is gratefully acknowledged.
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Published in the special issue Euroanalysis XVI (The European Conference on Analytical Chemistry) with guest editor Slavica Ražić.
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Serafín, V., Úbeda, N., Agüí, L. et al. Ultrasensitive determination of human growth hormone (hGH) with a disposable electrochemical magneto-immunosensor. Anal Bioanal Chem 403, 939–946 (2012). https://doi.org/10.1007/s00216-012-5753-9
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DOI: https://doi.org/10.1007/s00216-012-5753-9