Rapid determination of clenbuterol in urine by a competitive bead immunoassay based on Luminex technology.

A new competitive bead immunoassay (CBIA) based on Luminex technology for detecting clenbuterol in urine was reported. The carboxylated fluorescent beads were directly coated with clenbuterol derivatives without carrier protein spacer. Clenbuterol antibody was biotinylated, which was used for clenbuterol detection in combination with the functionalized bead and streptavidin-phycoerythrin (SAPE). The effects of spacer on the CBIA method were investigated. The results indicated that the presence of small molecular spacer between bead and hapten improved the assay sensitivity and the hydrophilic spacer (glycine) was better than the hydrophobic spacer (m-aminobenzoic acid) for this CBIA method. The study affirms the importance of the judicious choice of hapten derivatives in the CBIA method for detecting small molecule drug based on Luminex technology. The method could be used for clenbuterol detection in livestock urine and possible for the simultaneous detection of multiple veterinary drugs.

Development of an indirect enzyme-linked immunosorbent assay for the organophosphorus pesticide paraoxon-methyl.

In the present study, the synthesis of hapten for the organophosphorus (OP) pesticide paraoxon-methyl was developed, with a spacer arm (aminocarboxylic acid) attached at the aromatic ring. It was conjugated to bovine serum albumin (BSA) for use as an immunogen and to ovalbumin (OVA) for coating antigen for ELISA testing. Rabbits were immunized with the immunogen and two polyclonal antisera were produced and screened against the coating antigen using competitive indirect enzyme-linked immunosorbent assay (ELISA). For application to textile samples, the influence of several factors such as organic solvent, ionic strength, and pH on the ELISA results were studied. Under optimized conditions, the quantitative working range was 0.012-1.158 microg/mL with a limit of detection (LOD) of 0.005 microg/mL and the IC(50) was 0.115 microg/mL.There was negligible cross reactivity (CR) with other OP pesticides. The recoveries obtained by standard paraoxon-methyl addition to the different textile samples such as cotton, wool and muslin delaine were all from 86.0% to 108.0%. Therefore, the optimized ELISA may become a new convenient and economical analytical tool for monitoring paraoxon-methyl residues in textile samples.

Development of an immunochromatographic assay for rapid detection of 1-Aminohydantoin in urine specimens.

A rapid immunochromatographic assay was developed and validated for detection of 1-aminohydantoin (AHD) in urine specimens. Colloidal gold-labeled polyclonal antibody specific to AHD derivative was used as the marker; based on the competitive reactivity theory, the metabolite of nitrofurantoin after derivatization with benzaldehyde would compete with carboxyphenyl AHD derivative-conjugated ovalbumin. The test strip could efficaciously detect the novel analyte with a visual detection limit of 10 ng mL(-1) and high specificity. The reliability of the assay was determined by testing 80 standard samples comparing with enzyme-linked immunosorbent assay. The semi-quantitative detection was accomplished in less than 15 min with low cost, especially for requirements of rapid and simple screening. This is the first publication of an immunochromatographic assay for detection of nitrofuran residues.

Determination of medroxyprogesterone acetate residues by CE immunoassay with chemiluminescence detection.

A rapid and simple method is developed for the determination of medroxyprogesterone acetate (MPA) by CE immunoassay with chemiluminescence (CL). This method is based on the competitive reactions between horseradish peroxidase (HRP)-labeled MPA (MPA-HRP) and free MPA with anti-MPA antiserum. The influencing factors on the electrophoresis and CL detection were studied completely and the optimal conditions of separation and determination were obtained. The linear range was 2.0-50 nmol/L and the LOD for MPA was 0.9 nmol/L. The present method was applied to the analysis of pork tissues.

Development of a faster determination of 10 anabolic steroids residues in animal muscle tissues by liquid chromatography tandem mass spectrometry.

A method had been developed for determination of residues of 10 anabolic steroids (ASs) in animal muscle tissues by liquid chromatography tandem mass spectrometry (LC/MS/MS). After enzymolysis, the sample was extracted with tert-butyl methyl ether, cleaned up through reverse solid-phase extraction and further determined by LC/MS/MS under multiple reaction monitoring (MRM) mode. The limits of detection (LOD) of LC/MS/MS method used for testing epitestosterone (ETS), nandrolone (17 beta-NT), 17 alpha-methyl-testosterone (MTS), testosterone 17-propionate (PTS), medroxyprogesterone (MED), progesterone (PG), estrone (ESN), 17 beta-estradiol (17 beta-ES), 17alpha-ethynylestradiol (EES) and estriol (EST) in animal muscle ranged from 0.06 to 0.22 microg/kg, and the limits of quantification (LOQ) were from 0.12 to 0.54 microg/kg. Experiments on spiked samples of pork, beef, chicken and fish showed that at addition level of 1.0 microg/kg, the average recoveries of the ASs ranged from 64% to 77%, and coefficients of variation from 7.1% to 20.3%, while at addition level of 2.0 microg/kg, the average recoveries ranged from 70% to 89%, and coefficient of variation from 7.1% to 19.1%.