Development of an electrochemical ELISA for the screening of 17β-estradiol and application to bovine serum

Abstract

A sensitive electrochemical enzyme-linked immunosorbent assay (ELISA) for the detection of 17 beta-estradiol (17 beta-E2) was developed. Optimisation of two ELISA competition assays, using monoclonal or polyclonal antibodies anti-17 beta-estradiol, coupled with the electrochemical detection was firstly performed. The activity of the label enzyme (horseradish peroxidase) was measured electrochemically using 3,3',5,5'-tetramethylbenzidine as substrate. The use of the polyclonal antibody resulted in a more sensitive assay and the detection limit of the assay was estimated to be 20 pg ml-1. The analytical performances of the method were compared to those obtained using a dissociation enhanced lanthanide fluorescence immunoassay (DELFIA). Although sample extraction is not usually required by DELFIA, both extracted and non extracted samples were assayed. The comparison between the two screening techniques revealed similar results for the extracted samples and showed a comparable precision (RSD%), ranging from 6.2 to 13.4 and from 6.7 to 14.3 for DELFIA and ELISA, respectively. The results obtained by these screening assays were confirmed by liquid chromatography atmospheric pressure chemical ionisation tandem mass spectrometry which is currently used to confirm illegal hormone administration for regulatory purposes. The electrochemical enzyme immunoassay appears suitable as a screening tool for routine analysis of bovine serum estradiol and can be extended to other anabolic hormones using appropriate antibodies.

Photometric and Electrochemical Enzyme-Multiplied Assay Techniques Using β-Galactosidase as Reporter Enzyme

“…The response behavior, or dynamic range, is consistent with what is expected based upon antibody affinity to estradiol. There has been a report of an ELISA‐based electrochemical system for estradiol screening with lower detection limit capabilities; however, the homogeneous EMAT scheme avoids washing/separation steps and is more convenient (33).…”

Veterinary drug residues in foods

“…Various forms of chromatography are available for that purpose. Methods have been described for β -agonists (O'Keeffe et al 1999;Haasnoot et al 2002), chloramphenicol (Impens et al 2003;Scortichini et al 2005), nitro-imidazoles and anabolic steroids (Draisci et al 2000;Hageleit et al 2001), among others.…”

The Comamonas testosteroni steroid biosensor system (COSS)—Reflection on other methods

“…This method is based on the extent of competitive binding between sample estradiol and labelled estradiol to its specific capture antibody that has been immobilised on the immunosensor surface [27].…”