Objective To determine 3-amino-5-morpholinomethyl-2-oxazolidinone （AMOZ） residues released from protein bound AMOZ in animal tissues. Methods Polyclonal and monoclonal antibodies were produced in this study. A rapid...Objective To determine 3-amino-5-morpholinomethyl-2-oxazolidinone （AMOZ） residues released from protein bound AMOZ in animal tissues. Methods Polyclonal and monoclonal antibodies were produced in this study. A rapid, sensitive, and specific competitive direct enzyme-linked immunosorbent assay （cdELISA） was developed. Results Rabbit polyclonal antibodies were used in the optimized cdELISA method, and exhibited negligible cross-reactivity with other compounds structurally related to AMOZ. The IC50 of the polycional antibody was 0.16 ng/mL The method limit of detection in four different types of animal and fish tissues was less than 0.06 μg/kg. Recoveries ranged from 80% to 220% for fortified samples with the coefficient of variation values less than 15%. The results of the cdELISA method were in good agreement with the results from an established liquid chromatography-tandem mass spectrometry confirmatory method used for AMOZ residues. Conclusion The cdELISA method developed in the present study is a convenient practical tool for screening large numbers of animal and fish tissue samples for the the detection of released protein bound AMOZ residues.展开更多
3-氨基-5-吗啉甲基-2-恶唑烷酮(AMOZ)是抗生素呋喃它酮的代谢产物,具有潜在致畸、致癌性。由于分子量太小,其特异性抗体制备尚未突破,无法建立直接检测的免疫分析方法。为了制备针对AMOZ的特异性抗体,本研究首次尝试采用赖氨酸树状分子(...3-氨基-5-吗啉甲基-2-恶唑烷酮(AMOZ)是抗生素呋喃它酮的代谢产物,具有潜在致畸、致癌性。由于分子量太小,其特异性抗体制备尚未突破,无法建立直接检测的免疫分析方法。为了制备针对AMOZ的特异性抗体,本研究首次尝试采用赖氨酸树状分子(G5)替代传统载体蛋白,将AMOZ的乙醛酸衍生半抗原AMOZA直接偶联到G5分子上制备免疫原(AMOZA-G5),免疫Balb/c小鼠,收集抗血清并进行间接竞争酶联免疫分析(ci ELISA)测定。结果发现:传统免疫原AMOZA-BSA对应的抗血清无法特异性识别游离AMOZ,而AMOZA-G5对应的抗血清则可以特异性识别游离AMOZ,在ci ELISA实验中,1μg/m L AMOZ对抗体活性的抑制率为35%,说明以赖氨酸树状分子为载体可以有效提高小分子半抗原的免疫原性。本研究对于制备其他小分子半抗原的特异性抗体提供了方法借鉴。展开更多
基金supported by the National Science Foundation for Young Scientists of China(No.21107104)the State Key Program of National Natural Science of China(No.20837003)+1 种基金grants from the Ministry of Health(No.200902009)the National Science&Technology Pillar Program(No.2009BADB9B03-Z02)
文摘Objective To determine 3-amino-5-morpholinomethyl-2-oxazolidinone （AMOZ） residues released from protein bound AMOZ in animal tissues. Methods Polyclonal and monoclonal antibodies were produced in this study. A rapid, sensitive, and specific competitive direct enzyme-linked immunosorbent assay （cdELISA） was developed. Results Rabbit polyclonal antibodies were used in the optimized cdELISA method, and exhibited negligible cross-reactivity with other compounds structurally related to AMOZ. The IC50 of the polycional antibody was 0.16 ng/mL The method limit of detection in four different types of animal and fish tissues was less than 0.06 μg/kg. Recoveries ranged from 80% to 220% for fortified samples with the coefficient of variation values less than 15%. The results of the cdELISA method were in good agreement with the results from an established liquid chromatography-tandem mass spectrometry confirmatory method used for AMOZ residues. Conclusion The cdELISA method developed in the present study is a convenient practical tool for screening large numbers of animal and fish tissue samples for the the detection of released protein bound AMOZ residues.
文摘3-氨基-5-吗啉甲基-2-恶唑烷酮(AMOZ)是抗生素呋喃它酮的代谢产物,具有潜在致畸、致癌性。由于分子量太小,其特异性抗体制备尚未突破,无法建立直接检测的免疫分析方法。为了制备针对AMOZ的特异性抗体,本研究首次尝试采用赖氨酸树状分子(G5)替代传统载体蛋白,将AMOZ的乙醛酸衍生半抗原AMOZA直接偶联到G5分子上制备免疫原(AMOZA-G5),免疫Balb/c小鼠,收集抗血清并进行间接竞争酶联免疫分析(ci ELISA)测定。结果发现:传统免疫原AMOZA-BSA对应的抗血清无法特异性识别游离AMOZ,而AMOZA-G5对应的抗血清则可以特异性识别游离AMOZ,在ci ELISA实验中,1μg/m L AMOZ对抗体活性的抑制率为35%,说明以赖氨酸树状分子为载体可以有效提高小分子半抗原的免疫原性。本研究对于制备其他小分子半抗原的特异性抗体提供了方法借鉴。
