Characterization of polyclonal and monoclonal anti-taxol antibodies and measurement of taxol in serum.
Characterization of polyclonal and monoclonal anti-taxol antibodies and measurement of taxol in serum.
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发表时间：
1993-03
期刊：
影响因子：
11.2
通讯作者：
J. Leu;Bi-Xing Chen;Peter B. Schiff;Bernard F. Erlanger
中科院分区：
文献类型：
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作者：
J. Leu;Bi-Xing Chen;Peter B. Schiff;Bernard F. Erlanger
用2‘-琥珀酸基紫杉醇经混合酸化法制备的紫杉醇-牛血清白蛋白结合物在兔体内产生抗紫杉醇抗体。用2‘-琥珀酰紫杉醇-牛血清白蛋白免疫小鼠，可产生抗紫杉醇的多克隆抗体。用[~3H]紫杉醇进行放射免疫测定，标准曲线显示其半数抑制浓度为1.0 nM。人血清中紫杉醇的检测下限为0.1nM或0.085 ng/ml，用相同抗原免疫BALB/c小鼠，可获得两株小鼠抗紫杉醇单抗。一种为免疫球蛋白G1(69E4A8E)，另一种为免疫球蛋白M(29B7B3C)。通过与紫杉醇和10种不同的相关衍生物及其类似物的竞争酶联免疫吸附试验来确定这些抗体的特异性。29B7B3C对生物活性衍生物具有较高的结合亲和力，而对非活性衍生物的亲和力明显较低，即其特异性与相同紫杉醇衍生物的微管蛋白拆解和细胞毒性研究结果一致，适合于天然产物提取物中紫杉醇或紫杉醇类化合物的筛选。69E4A8E识别紫杉醇C-3‘位的苯甲酰胺氨基甲酰基，与不同取代基的活性化合物亲和力较低。人血清中紫杉醇水平可用69E4A8E竞争酶联免疫吸附测定法进行检测。测定下限约为50 nM或约42 ng/ml，可用放射免疫法测定。
Anti-taxol antibodies were generated in the rabbit using a taxol-bovine serum albumin conjugate prepared from 2'-succinyltaxol using a mixed anhydride procedure. Immunization with 2'-succinyltaxol-bovine serum albumin gave rise to polyclonal anti-taxol antibodies. By a radioimmunoassay using [3H]taxol, a standard curve gave a 50% inhibitory concentration of 1.0 nM. Taxol levels in human serum could be measured, with the lower limit of detection and measurement being 0.1 nM or 0.085 ng/ml. Two mouse monoclonal anti-taxol antibodies were isolated by immunizing BALB/c mice with the same antigen. One was an immunoglobulin G1 (69E4A8E) and the other was immunoglobulin M (29B7B3C). The specificity of these antibodies was determined by a competitive enzyme-linked immunosorbent assay with taxol and 10 different related derivatives and analogues. 29B7B3C had higher binding affinities for biologically active derivatives and markedly lower affinities for inactive derivatives; i.e., the specificity was consistent with the results of tubulin disassembly and cytotoxicity studies using the same taxol derivatives, making it suitable for screening for taxol or taxol-like compounds in extracts of natural products. 69E4A8E recognized the benzamidocarbamyl group at the C-3' position of taxol and had a lower affinity for other active compounds with different substitutions. Taxol levels in human serum could be detected and measured by 69E4A8E using a competitive enzyme-linked immunosorbent assay. The lower limit of measurement was about 50 nM or approximately 42 ng/ml. Similar measurements could be made by radioimmunoassay.