Enzyme immunoassay (ELISA/immunosensor) for a sensitive detection of pork adulteration in meat.

ELISA/immunosensor were developed for a sensitive detection of pork adulteration in meat. Two formats of ELISA were performed. First, an extracted IgG was directly immobilized in the microplate. This assay allowed an identification of 0.01% as level of pork adulteration in 14 h15 min. In order to decrease the time of the assay, a competitive ELISA was developed by immobilizing IgG standard, which compete with the extracted IgG. This assay allowed a detection of 0.1% of pork adulteration in 45 min. Furthermore, two formats of electrochemical immunosensors were elaborated using the electro-entrapment of IgG in polymer modified graphite paste electrode. First, a direct immunosensor was capable of identifying 0.1 and 1% in raw and cooked meat respectively in 2 h. The second format was based on a competitive immunosensor, which was able to detect 0.01% of pork adulteration within 20 min. Both competitive immunoassays revealed high sensitivity, good specify and reduced analysis time.

Electrochemical DNA sandwich biosensor based on enzyme amplified microRNA-21 detection and gold nanoparticles.

In this work, a novel electrochemical biosensor for miRNA-21 determination, involving a sandwich hybridization assay onto gold nanoparticles modified pencil graphite electrode (PGE) and enzyme signal amplification was reported. The thiol terminated capture probe 1 (SH-P1) was immobilized on the electrode through AuS interaction. In the presence of target miRNA-21, SH-P1 hybridized with the first part of the target, however, the second part hybridizes with a biotinylated probe P2 (B-P2). Then, a streptavidin-conjugated alkaline phosphatase was immobilized by a specific binding of avidin-B-P2. The enzyme catalyzed the electro-inactive α-naphtyl phosphate to an electro-active α-naphtol. The miRNA-21 detection was achieved through the changes of α-naphtol oxidation signals observed at +0.12V vs Ag/AgCl with Differential Pulse Voltammetry. Under the optimal detection conditions, the biosensor exhibited selective and sensitive detection with a linear range from 200pM to 388nM and the detection limit was 100pM (10fmol in 100µL).

A novel method for sensitive microRNA detection: Electropolymerization based doping.

In the proposed study, for the first time, sensitive electrochemical detection of a breast cancer biomarker microRNA (miRNA), mir-21 was achieved via electropolymerized polypyrrole (PPy) modified pencil graphite electrodes (PPy/PGE). The detection of hybridization of electrochemically doped probe miRNA, antimir-21, with its complementary target, mir-21 was monitored by either electrochemical impedance spectroscopy (EIS) via comparison of charge transfer resistance (Rct) values before and after hybridization or by electrochemical reduction signal of an hybridization indicator, Meldola's blue (MDB). The study covers all the optimization steps for hybridization procedure and electropolymerization of pyrrole as well as detection from real samples of breast cancer cell line, MCF-7. The designed sensor shows a high selectivity and a low detection limit of 0.17nM thanks to electrical conductivity and porous structure of PPy.