An immunoassay for dibutyl phthalate based on direct hapten linkage to the polystyrene surface of microtiter plates.
An immunoassay for dibutyl phthalate based on direct hapten linkage to the polystyrene surface of microtiter plates.
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基于半抗原与微量滴定板聚苯乙烯表面的直接连接的邻苯二甲酸二丁酯的免疫测定。
DOI：
10.1371/journal.pone.0029196
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发表时间：
2011
期刊：
影响因子：
3.7
通讯作者：
Yuan J
中科院分区：
文献类型：
--
作者：
Wei C;Ding S;You H;Zhang Y;Wang Y;Yang X;Yuan J
邻苯二甲酸二丁酯 (DBP) 主要用作塑料中的增塑剂，以使其具有柔韧性。邻苯二甲酸盐在工业过程和其他消费品中的广泛使用导致邻苯二甲酸盐在环境中无处不在。为了更好地确定环境中的污染程度并评估暴露于DBP的潜在不利影响，开发了DBP免疫测定法。 DBP 特异性单克隆抗体是由淋巴细胞杂交瘤技术产生的稳定杂交瘤细胞系产生的。建立了一种直接在聚苯乙烯微量滴定板上涂覆半抗原的间接竞争酶联免疫吸附测定（icELISA）来检测 DBP。聚苯乙烯表面首先在稀硫酸中被高锰酸盐氧化生成羧基。然后将 DBP 类似物 4-氨基邻苯二甲酸二丁酯与 1-乙基-3-(3-二甲基氨基丙基)碳二亚胺盐酸盐 (EDC) 共价连接到聚苯乙烯表面的羧基上。与缀合物包被形式 (IC50= 106 ng/mL) 相比，直接半抗原包被形式 (IC50= 14.6 ng/mL) 在仔细优化测定条件后提高了测定灵敏度。加标水样中DBP的平均回收率为104.4%，平均变异系数为9.95%。半抗原包被的 icELISA 和气相色谱-质谱法获得的结果非常吻合，进一步证实了 icELISA 用于检测某些塑料和化妆品样品中 DBP 的可靠性和准确性。获得的稳定、高效的杂交瘤细胞系是敏感、特异的DBP抗体的无限来源。半抗原包被的形式被认为是普遍适用的，因为修饰的微量滴定板表面上的羧基能够使胺化或羟基化半抗原与EDC稳定固定。开发的半抗原包被的 icELISA 可作为一种方便的定量工具，用于灵敏、准确地监测水、塑料和化妆品样品中的 DBP。
Dibutyl phthalate (DBP) is predominantly used as a plasticizer inplastics to make them flexible. Extensive use of phthalates in both industrial processes and other consumer products has resulted in the ubiquitous presence of phthalates in the environment. In order to better determine the level of pollution in the environment and evaluate the potential adverse effects of exposure to DBP, immunoassay for DBP was developed. A monoclonal antibody specific to DBP was produced from a stable hybridoma cell line generated by lymphocyte hybridoma technique. An indirect competitive enzyme-linked immunosorbent assay (icELISA) employing direct coating of hapten on polystyrene microtiter plates was established for the detection of DBP. Polystyrene surface was first oxidized by permanganate in dilute sulfuric acid to generate carboxyl groups. Then dibutyl 4-aminophthalate, which is an analogue of DBP, was covalently linked to the carboxyl groups of polystyrene surface with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC). Compared with conjugate coated format (IC50 = 106 ng/mL), the direct hapten coated format (IC50 = 14.6 ng/mL) improved assay sensitivity after careful optimization of assay conditions. The average recovery of DBP from spiked water sample was 104.4% and the average coefficient of variation was 9.95%. Good agreement of the results obtained by the hapten coated icELISA and gas chromatography-mass spectrometry further confirmed the reliability and accuracy of the icELISA for the detection of DBP in certain plastic and cosmetic samples. The stable and efficient hybridoma cell line obtained is an unlimited source of sensitive and specific antibody to DBP. The hapten coated format is proposed as generally applicable because the carboxyl groups on modified microtiter plate surface enables stable immobilization of aminated or hydroxylated hapten with EDC. The developed hapten coated icELISA can be used as a convenient quantitative tool for the sensitive and accurate monitoring DBP in water, plastic and cosmetic samples.
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影响因子：
7.4
作者：
Estévez, MC;Galve, R;Marco, MP
通讯作者：
Marco, MP
影响因子：
6.1
作者：
Krämer, PM;Goodrow, MH;Kremmer, E
通讯作者：
Kremmer, E
影响因子：
6.1
作者：
Ramon-Azcon, Javier;Sanchez-Baeza, Francisco;Marco, M. -Pilar
通讯作者：
Marco, M. -Pilar
影响因子：
44.1
作者：
Hu, YX;Guo, JY;Zhang, YL
通讯作者：
Zhang, YL
影响因子：
--
作者：
TEMPONI, M;KAGESHITA, T;FERRONE, S
通讯作者：
FERRONE, S