Neamin as an Immunogen for the Development of a Generic ELISA Detecting Gentamicin, Kanamycin, and Neomycin in Milk

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• 作者：Elma E. M. G. Loomans ; Jim van Wiltenburg ; Marjo Koets ; and Aart van Amerongen
• 刊名：Journal of Agricultural and Food Chemistry
• 出版年：2003
• 出版时间：January 29, 2003
• 年：2003
• 卷：51
• 期：3
• 页码：587 - 593
• 全文大小：117K
• 年卷期：v.51,no.3(January 29, 2003)
• ISSN：1520-5118

文摘

A broad-specific ELISA using one antibody preparation for the detection of gentamicin, kanamycin,and neomycin in milk is reported for the first time. For the immunization of rabbits, neamin was usedas the generic hapten on the basis of the facts that it is a two-ring fragment of neomycin and, inshape and charge, it resembles parts of kanamycin and gentamicin. Neamin was linked to thepreactivated carrier protein keyhole limpet hemocyanin by EDC coupling. The specificity and sensitivityof the polyclonal antibodies for the aminoglycoside antibiotics were tested in a competitive assayusing homologous and heterologous conjugates coupled by various conjugation procedures as theELISA solid phase. In contrast to the homologous assay recognizing only neomycin, the heterologousassay could be used for the detection of the whole subclass of deoxystreptamin antibiotics in bufferand raw milk. Gentamicin, kanamycin, and neomcyin were detected in artificially contaminatedundiluted raw milk (without sample pretreatment) with 50% inhibition levels at 9, 21, and 113 ngmL^-1, respectively. Neomycin levels were also measured in milk samples obtained from a cow sufferingfrom mastitis and treated with an antibiotic cocktail including neomycin. Levels below the EU maximumresidue levels for deoxystreptamin antibiotics could readily be measured in this generic ELISA.Keywords: Antibiotic; aminoglycoside; gentamicin; kanamycin; neomycin; dihydrostreptomycin;neamin; generic hapten; generic antibody; ELISA; milk