Clenbuterol immunoassay

Elliott et al. utilized a clenbuterol immunoassay to determine clenbuterol residues in cattle tissues and fluids. The LOD was 0.25 ug for liver. Animals were dosed with medicated feed (1.6 ug kg per day), and pairs were slaughtered during the medication phase and at 14,28, and 42 days after withdrawal. Clenbuterol concentrations in liver and retina/choroid samples were confirmed by GC/MS. Correlation coefficients between the ELISA and GC/MS were = 0.92 for retina/choroid samples and... 

Shelver and Smittf confirmed fhaf commercial clenbuterol immunoassays cross-react with some, but not all, clenbuterol metabolites. As a result, quantitative clenbuterol immunoassays may differ from determinative methods if substantial concentrations of metabolites are present. For clenbuterol, the parent clenbuterol level is... 

Matsumoto et al. developed an immunoassay for the determination of clenbuterol in bovine and equine tissues and in bovine milk. The LOD of clenbuterol in milk, muscle, liver, kidney, small intestine, and adipose tissues was 0.1 qgkg Bovine tissue samples fortified wifh 1 qg kg of clenbuterol had recoveries that varied from 75 to 96%, but recoveries from milk samples were 99%. The authors utilized this method to estimate the clenbuterol withdrawal periods for cattle and horses. Cattle were treated with a bolus dose of either 0.3 or 0.6 qg kg body weight, by intravenous injection, and three animals were slaughtered at days 1, 6, and 9. Tissue clenbuterol levels were detectable only on day 1. Clenbuterol in milk was not detectable after a 2.5-day withdrawal period. Liver contained the highest clenbuterol concentration of the tissues measured, but this group did not measure eye tissues. 

To test our new signal reagent based on GZ-11 the detection system was applied to two competitive-format immunoassays. These two assays (for atrazine and clenbuterol) normally use chromogenic detection systems. While these colorimetric assays may be adequate for laboratory use, chemiluminescent detection offers potential advantages in sensitivity and on site screening applications [33],... 

Optimization of the immunoassay was performed with respect to tracer and antibody concentrations to obtain the required sensitivity. These conditions differed depending on the detection system used photographic detection required higher antibody and tracer concentrations than when the plate luminometer was used. A further complication arose from the very low affinity of the tracer for the antibody when using an antibody dilution of 1 3000 and a tracer dilution of 1 4000 less than 1% of the tracer was bound after a 2-h incubation. This means that the antibody, in the absence of clenbuterol, binds less than 10 pg of the... 

Examples exist of different types of immunobased technology being interfaced to produce an effective analytical system. In a variety of recent methods, immunoaffinity chromatography has been employed for purification, chemiluminescence enzyme immunoassay has been used for quantification of salbutamol and clenbuterol in tissue and plasma from calves and pigs (122), clenbuterol in cattle hair (123), and monensin in chicken tissues (124). In these methods, quantification at sub-ppb levels has been demonstrated. 

In 1982, the first enzyme immunoassay of clenbuterol was described (134). It was used to determine clenbuterol levels in plasma of human patients treated by oral route with this drug. It was a highly sensitive double-antibody and heterologous immunoassay based on a competition for binding to a clenbuterol-specific antibody between a diazotized clenbuterol analogue labeled with -galactosidase and unlabeled standard or sample clenbuterol. The antibody-bound enzyme hapten was separated from free hapten by anti-rabbit IgG immobilized to a polystyrene ball. The assay could detect levels as low as 0.5 pg clenbuterol per tube. 

Instead of using human serum albumin as a carrier protein, other workers (135) utilized ovalbumin for preparing the diazotized clenbuterol antigen in an enzyme immunoassay developed for screening of clenbuterol residues in bovine urine, liver, and eye. Alkaline phosphatase rather than -galactosidase was also used as an enzyme label in the preparation of the enzyme-clenbuterol conjugate. 

Unlike clenbuterol, salbutamol is a difficult compound to analyze due to its particular chemical attributes. It is a basic compound subjected to protein binding poor recoveries are obtained especially when protein precipitation techniques are used to prepare the extracts (145). In addition, salbutamol is charged at all pH values and does not readily lend itself to simple, specific back-extracting procedures. This severely restricts the options of sample cleanup. However, a Subtilisin protease digestion step followed by acid clarification and solid-phase extraction has been suggested (146) as an adequate extraction and cleanup procedure prior to the end-point determination of salbutamol by an enzyme immunoassay (139) based on the cross-reactivity of anticlenbuterol antibodies. 

G Degand, et al. Determination of clenbuterol in bovine tissues and urine by enzyme immunoassay. J Agric Food Chem 40 70, 1992. 

RJH Picket, MJ Sauer. Determination of clenbuterol in bovine urine by enzyme immunoassay following concentration and clean-up by immunoaffinity chromatography. Anal Chim Acta 275 268, 1993. 

M.E. Ploum, W. Haasnoot, R.J.A. Paulussen, G.D. Van Bruchem, A. Hames, R. Schilt, F.A. Hug, Test Strip Enzyme Immunoassays and the Fast Screening of Nortesterone and Clenbuterol Residues in Urine Samples at the Parts per Billion Level , I. Chromatogr., 546, 413-427 (1991). 

Lai, Y., Bai, J., Shi, X. et al. (2013) Graphene oxide as nanocarrier for sensitive electrochemical immunoassay of clenbuterol based on labeling amplification strategy. Talanta, 107, 176-182. 

Bai, J., Lai, Y, Jiang, D. et al. (2012) Ultrasensitive electrochemical immunoassay based on graphene oxide-Ag composites for rapid determination of clenbuterol Analyst, 137,4349-4355. 

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