Rapid determination of recent cocaine use with magnetic particles-based enzyme immunoassays in serum, saliva, and urine fluids.

Cocaine is one of the most worldwide used illicit drugs. We report a magnetic particles-based enzyme-linked immunoassay (mpEIA) method for the rapid and sensitive determination of cocaine (COC) in saliva, urine and serum samples. Under optimized conditions, the limits of detections were 0.09ngmL(-1) (urine), 0.15ngmL(-1) (saliva), and 0.06ngmL(-1) COC (human serum). Sensitivities were in the range EC50=0.6-2.5ngmL(-1) COC. The cross-reactivity with the principal metabolite benzoylecgonine (BZE) was only 1.6%. Recovering percentages of doped samples (0, 10, 50, and 100ngmL(-1) of COC) ranged from about 86-111%. Some advantages of the developed mpEIA over conventional ELISA kits are faster incubations, improved reproducibility, and consumption of lower amounts of antibody and enzyme conjugates due to the use of magnetic beads. The reported method was validated following the guidelines on bioanalytical methods of the European Medicines Agency (2011). Unmetabolized COC detection has a great interest in pharmacological, pharmacokinetics, and toxicokinetics studies, and can be used to detect a very recent COC use (1-6h).

Electrochemical affinity biosensors for detection of mycotoxins: A review.

This review discusses the current state of electrochemical biosensors in the determination of mycotoxins in foods. Mycotoxins are highly toxic secondary metabolites produced by molds. The acute toxicity of these results in serious human and animal health problems, although it has been only since early 1960s when the first studied aflatoxins were found to be carcinogenic. Mycotoxins affect a broad range of agricultural products, most important cereals and cereal-based foods. A majority of countries, mentioning especially the European Union, have established preventive programs to control contamination and strict laws of the permitted levels in foods. Official methods of analysis of mycotoxins normally requires sophisticated instrumentation, e.g. liquid chromatography with fluorescence or mass detectors, combined with extraction procedures for sample preparation. For about sixteen years, the use of simpler and faster analytical procedures based on affinity biosensors has emerged in scientific literature as a very promising alternative, particularly electrochemical (i.e., amperometric, impedance, potentiometric or conductimetric) affinity biosensors due to their simplicity and sensitivity. Typically, electrochemical biosensors for mycotoxins use specific antibodies or aptamers as affinity ligands, although recombinant antibodies, artificial receptors and molecular imprinted polymers show potential utility. This article deals with recent advances in electrochemical affinity biosensors for mycotoxins and covers complete literature from the first reports about sixteen years ago.

An electrochemical immunosensor for ochratoxin A determination in wines based on a monoclonal antibody and paramagnetic microbeads.

We report a direct competitive immunosensor for the rapid determination of ochratoxin A (OTA) in wine samples. Magnetic beads (1 ± 0.5 µm diameter) covered with streptavidin were functionalized with a monoclonal antibody against OTA, and then left to incubate in a solution of tracer (ochratoxin conjugated to the enzyme peroxidase) and a range of OTA concentrations (10(-4) to 1,000 ng mL(-1)). After washing and separation steps helped with a magnetic field, a volume of the dispersion was put on screen-printed electrodes under a magnet, and after adding the substrate the p-benzoquinone generated enzymatically was detected by differential-pulse voltammetry. Wine samples (2 mL) were easily prepared simply by adjusting to pH = 7.5 with diluted NaOH and by adding polyvinylpyrrolidone for complexing polyphenols, without any other clean-up or preconcentration steps. The limit of detection for detecting OTA in wines was of 0.11 ± 0.01 ng L(-1), well below the permitted content of the mycotoxin by the European Union (<2 ng mL(-1)). Spiked wines were subjected to immunosensor calibrations to study the matrix effects. OTA concentrations measured with the immunosensor were compared with those obtained by high-performance liquid chromatography coupled to fluorescence detection (AOAC official method 2001.01). The OTA levels from two red wines of "Campo de Borja", Spain, ranged from about 0.027 to 0.033 ng mL(-1) of OTA.

An electrochemical competitive biosensor for ochratoxin A based on a DNA biotinylated aptamer.

Ochratoxin A (OTA) is one of the most important mycotoxin contaminants of foods, particularly cereals and cereal products, with strict low regulatory levels (of ppb) in many countries worldwide. An electrochemical competitive aptamer-based biosensor for OTA is described. Paramagnetic microparticle beads (MBs) were functionalized with an aptamer specific to OTA, and were allowed to compete with a solution of the mycotoxin conjugated to the enzyme horseradish peroxidase (OTA-HRP) and free OTA. After separation and washing steps helped with magnetic separations, the modified MBs were localized on disposable screen-printed carbon electrodes (SPCEs) under a magnetic field, and the product of the enzymatic reaction with the substrate was detected with differential-pulse voltammetry. In addition to magnetic separation assays, other competitive schemes (direct/indirect aptasensors performed on the SPCEs surface or using gold nanoparticles functionalized with the aptamer) were preliminary tested, optimized and compared. The magnetic aptasensor showed a linear response to OTA in the range 0.78-8.74 ng mL(-1) and a limit of detection of 0.07±0.01 ng mL(-1), and was accurately applied to extracts of certified and spiked wheat samples with an RSD lower than about 8%.