Development of an immunoassay for the beta-adrenergic agonist ractopamine.
Antibody generated from ractopamine-hemiglutarate-KLH was used to develop a ractopamine ELISA. The antibody showed good sensitivity in phosphate buffer, with an IC50 of 4.2 ng/ml (ppb) toward ractopamine and 16.2 ng/ml toward glucuronides of ractopamine conjugated to the phenethanolamine phenol of ractopamine. Phenylbutylamine phenol glucuronides of the (RS, SR) ractopamine diastereoisomers showed about 4% cross-reactivity, but the glucuronide of the (RR, SS) diastereoisomers conjugated at the same phenolic group showed no detectable reactivity with the antibody. The antibody generally had cross-reactivity towards compounds with bis-phenylalkyl amine structures rather than compounds with simple branched N-alkyl substituents. For example, the antibody showed little or no cross reactivity towards clenbuterol, isoproterenol, metaproterenol, and salbutamol, but cross-reacted with dobutamine. The system demonstrated a matrix effect similar to other enzyme immunoassays, dilution of urine decreased but did not eliminate the matrix effect.
Shelver
W L
WL
US Department of Agriculture, Biosciences Research Laboratory, University Station, Fargo, ND 58105-5674, USA. shelverw@fargo.usda.ars.gov
Smith
D J
DJ
D000318:Adrenergic beta-Agonists / Q000032:analysis* / Q000737:chemistry / Q000276:immunology / Q000493:pharmacokinetics; D000812:Anhydrides / Q000737:chemistry; D000818:Animals; D000906:Antibodies / Q000302:isolation & purification / Q000378:metabolism; D000918:Antibody Specificity; D001667:Binding, Competitive; D002417:Cattle; D002846:Chromatography, Affinity; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005069:Evaluation Studies as Topic; D020719:Glucuronides / Q000138:chemical synthesis / Q000737:chemistry / Q000652:urine; D005977:Glutarates / Q000737:chemistry; D006241:Haptens / Q000737:chemistry / Q000276:immunology; D006433:Hemocyanins / Q000737:chemistry / Q000276:immunology; D010627:Phenethylamines / Q000032:analysis* / Q000737:chemistry / Q000276:immunology / Q000493:pharmacokinetics; D011817:Rabbits; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D000318:Adrenergic beta-Agonists; D000812:Anhydrides; D000906:Antibodies; D020719:Glucuronides; D005977:Glutarates; D006241:Haptens; D010627:Phenethylamines; C060897:ractopamine; C079643:glutaric anhydride; D006433:Hemocyanins; C032808:keyhole-limpet hemocyanin