Nussberger J, Keller I, Waeber B, Brunner H R
Division of Hypertension, University Hospital, Lausanne, Switzerland.
J Hypertens Suppl. 1988 Dec;6(4):S424-5. doi: 10.1097/00004872-198812040-00133.
Selectivity for the carboxy-terminus of angiotensin II (Ang II) and the high affinity of antibodies are prerequisites for clinical assays that evaluate Ang II in the presence of Ang I. A high-affinity monoclonal antibody (Kd = 7.1 X 10(-11) mol/l) was produced and used for the measurement of plasma Ang II. C3H mice were immunized with Ang II coupled by its carboxy-terminus to thyroglobulin. Somatic cell fusion between spleen cells and SP 2/0 myeloma cells, repeated subcloning and re-injection into mice yielded ascites containing sufficient antibody at a 2 X 10(7)-fold dilution. Radioassay standard curves show 50% tracer displacement when 32 fmol unlabelled Ang II is added and 2 fmol Ang II can be detected. Cross-reactivities, taking the reactivity with Ang II as 1.00 are: Ang I 0.003, Ang (1-7) 0.00001, Ang III 1.05, Ang(3-8) 0.88 and Ang(4-8) 0.75. Fast extraction of angiotensin from 2 ml plasma by reversible adsorption to phenylsilylsilica (Bondelut PH) provides recoveries of 96-102%. During angiotensin converting enzyme inhibition with 25 mg intravenous captopril, plasma immunoreactive Ang II decreased in supine normal volunteers from 8.6 +/- 3.6 to 4.5 +/- 3.4 fmol/ml (P less than 0.01, n = 8). It thus appears that plasma immunoreactive Ang II can now be measured after a simple extraction procedure by using monoclonal antibodies.
对血管紧张素II（Ang II）羧基末端的选择性以及抗体的高亲和力是在存在血管紧张素I（Ang I）的情况下评估Ang II的临床检测的先决条件。制备了一种高亲和力单克隆抗体（解离常数Kd = 7.1×10⁻¹¹mol/L）并用于测量血浆Ang II。用通过其羧基末端与甲状腺球蛋白偶联的Ang II免疫C3H小鼠。脾细胞与SP 2/0骨髓瘤细胞进行体细胞融合，反复亚克隆并重新注射到小鼠体内，产生腹水，腹水中含有足够的抗体，稀释倍数为2×10⁷倍。放射分析标准曲线显示，加入32 fmol未标记的Ang II时，示踪剂位移50%，可检测到2 fmol Ang II。以与Ang II的反应性为1.00，交叉反应性如下：Ang I为0.003，Ang (1 - 7)为0.00001，Ang III为1.05，Ang(3 - 8)为0.88，Ang(4 - 8)为0.75。通过可逆吸附到苯基硅基硅胶（Bondelut PH）上从2 ml血浆中快速提取血管紧张素，回收率为96 - 102%。在25 mg静脉注射卡托普利抑制血管紧张素转换酶期间，仰卧位正常志愿者的血浆免疫反应性Ang II从8.6±3.6降至4.5±3.4 fmol/ml（P＜0.01，n = 8）。因此，现在似乎可以通过简单的提取程序后使用单克隆抗体来测量血浆免疫反应性Ang II。
