Mizusawa E, Dahlman H L, Bennett S J, Goldenberg D M, Hawthorne M F
Proc Natl Acad Sci U S A. 1982 May;79(9):3011-4. doi: 10.1073/pnas.79.9.3011.
Two samples of 2-phenyl-1,2-dicarba-closo-[1-3H]dodecaborane(12) were prepared by treating 1-lithio-2-phenyl-1,2-dicarba-closo-dodecaborane(12) with 3H2O (0.1 and 5.0 Ci/ml, respectively). These tritiated phenylcarborane samples were subsequently converted to corresponding samples of p-[1,2-dicarba-closo-[1-3H]dodecaboran(12)-2-yl]benzenediazonium ion ([3H]DBD) suitable for azo-coupling reactions. Reaction of the two tritiated diazonium ion samples with 2-napthol resulted in the formation of an azo dye (epsilon = 1.98 X 10(4) M-1 cm-1 at 485 nm). Experiments relating absorbance to 3H activity proved the two [3H]DBD sources to have 3.81 X 10(11) and 2.45 X 10(13) cpm of 3H per mol of tritiated carborane substituent. Purified antibodies to carcinoembryonic antigen were coupled to the [3H]DBD and, after extensive dialysis, the average number of carborane moieties per antibody molecule was determined by measuring the 3H activity associated with a known protein concentration. Further examination of these tritiated carborane-labeled antibodies by affinity chromatography proved that boron labeling did not destroy their immunoreactivity. Correlations of azo-coupling conditions (reactant ratios, pH) with immunoreactivity and antibody protein recovery are presented.
通过用3H2O（分别为0.1和5.0 Ci/ml）处理1-锂代-2-苯基-1,2-二碳杂-闭式-十二硼烷（12），制备了两份2-苯基-1,2-二碳杂-闭式-[1-3H]十二硼烷（12）样品。随后，将这些氚代苯基碳硼烷样品转化为适合偶氮偶联反应的对-[1,2-二碳杂-闭式-[1-3H]十二硼烷（12）-2-基]苯重氮离子（[3H]DBD）的相应样品。两份氚代重氮离子样品与2-萘酚反应生成了一种偶氮染料（在485 nm处ε = 1.98×10(4) M-1 cm-1）。将吸光度与3H活性相关的实验证明，每摩尔氚代碳硼烷取代基，两种[3H]DBD源的3H活度分别为3.81×10(11)和2.45×10(13) cpm。将癌胚抗原的纯化抗体与[3H]DBD偶联，经过广泛透析后，通过测量与已知蛋白质浓度相关的3H活性，确定每个抗体分子的碳硼烷部分的平均数量。通过亲和色谱对这些氚代碳硼烷标记的抗体进行进一步检测，证明硼标记不会破坏它们的免疫反应性。本文还介绍了偶氮偶联条件（反应物比例、pH）与免疫反应性和抗体蛋白回收率之间的相关性。
