Radioimmunoassay of the leukotrienes of slow reacting substance of anaphylaxis.
Radioimmunoassay of the leukotrienes of slow reacting substance of anaphylaxis.
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过敏反应慢反应物质白三烯的放射免疫测定。
DOI：
10.1073/pnas.78.12.7692
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发表时间：
1981
影响因子：
11.1
通讯作者：
Corey,EJ
中科院分区：
文献类型：
--
作者：
Levine,L;Morgan,RA;Lewis,RA;Austen,KF;Clark,DA;Marfat,A;Corey,EJ
通过二十烷酸羧基用白三烯 D4 (LTD4) 和牛白蛋白的缀合物免疫的兔子产生了对白三烯 C4、D4 和 E4（LTC4、LTD4 和 LTE4）及其 11-反式立体异构体具有相当亲和力的抗体。抗体在 37℃ 下以 2.8 x 10(9) M-1 的相同平均缔合常数 (Ka) 结合 3H 标记的 11-trans-LTC4 和 11-trans-LTC4，并且以 0.32 微克/ml 免疫兔血浆的浓度存在。当 9.5 微升抗 LTD4 和 108 pmol 11-反式-[3H]LTC4 (40 Ci/mmol) 在 300 微升体积中与 LTC4、LTD4、LTE4 或其 11-反式立体异构体一起孵育时，在 0.3 至 0.7 ng 之间的水平下实现了 11-反式-[3H]LTC4 结合的 50% 抑制。使用天然白三烯的合成类似物进行评估时，抗体既不识别由脱氨基或适度肽延长产生的 LTD4 6-硫肽单元内的那些变化，也不识别 δ 14-顺式双键的特定立体化学。然而，抗体确实识别三烯脂质结构域以及谷胱甘肽或半胱氨酰甘氨酸功能的位置和空间方向。抗 LTD4 与 11-反式-[3H]LTC4 的结合不受谷胱甘肽、胱氨酸双甘氨酸、花生四烯酸或 5-羟基-6,8,11,14-二十碳四烯酸的抑制，并且白三烯 B4 (LTB4) 的活性约为 LTC4、LTD4 或 LTE4 的 1/1000。小鼠淋巴瘤 (WEHI-5) 和大鼠嗜碱性粒细胞白血病 (RBL-1) 细胞在用钙离子载体 A23187 刺激时，各自产生免疫反应性白三烯； RBL-1 细胞中的 LTC4、LTD4 和 LTE4 在高效液相色谱解析后通过放射免疫测定法单独定量。
A rabbit immunized with a conjugate of leukotriene D4 (LTD4) and bovine albumin via the icosanoid carboxyl produced antibodies with comparable affinities for leukotrienes C4, D4, and E4 (LTC4, LTD4, and LTE4) and their 11-trans stereoisomers. The antibodies bound 3H-labeled 11-trans-LTC4 and 11-trans-LTC4 with the same average association constant (Ka) of 2.8 x 10(9) M-1 at 37 degrees C and were present at a concentration of 0.32 microgram/ml of the immune rabbit plasma. When 9.5 microliter of anti-LTD4 and 108 pmol of 11-trans-[3H]LTC4 (40 Ci/mmol) were incubated in a volume of 300 microliter with LTC4, LTD4, LTE4, or their 11-trans stereoisomers, 50% inhibition of 11-trans-[3H]LTC4 binding was achieved at levels varying between 0.3 and 0.7 ng. As assessed with synthetic analogs of the natural leukotrienes, the antibodies recognized neither those changes within the 6-sulfidopeptide unit of LTD4 produced by deamination or modest peptide lengthening nor the specific stereochemistry of the delta 14-cis double bond. However, the antibodies did recognize the triene lipid domain and the position and spatial orientation of the glutathione or cysteinylglycine function. Binding of 11-trans-[3H]LTC4 by anti-LTD4 was not inhibited by glutathione, cystinylbisglycine, arachidonic acid, or 5-hydroxy-6,8,11,14-icosatetraenoic acid, and leukotriene B4 (LTB4) was about 1/1000th as active as LTC4, LTD4, or LTE4. Mouse lymphoma (WEHI-5) and rat basophil leukemia (RBL-1) cells, when stimulated with calcium ionophore A23187, each produced immunoreactive leukotrienes; and LTC4, LTD4, and LTE4 from RBL-1 cells were individually quantitated by radioimmunoassay after resolution by high-performance liquid chromatography.