Korsching S, Thoenen H
Proc Natl Acad Sci U S A. 1983 Jun;80(11):3513-6. doi: 10.1073/pnas.80.11.3513.
A two-site enzyme immunoassay is described which does not suffer from artifacts inherent in previous assays and has the necessary high sensitivity to determine the endogenous levels of nerve growth factor (NGF) in the sympathetic nervous system and its target organs. Monoclonal and affinity-purified polyclonal antibodies against mouse NGF (mNGF) were covalently linked to glass beads as the first site and coupled to the enzyme beta-galactosidase as the second site. Detection of the fluorescent beta-galactosidase reaction product permitted the determination of 0.01-0.02 fmol of mNGF per assay. The recovery of mNGF added to homogenates varied between 50% and 100%, depending on the tissue. Rat superior cervical and stellate ganglia were found to contain (mean +/- SEM) 25 +/- 4 and 19 +/- 3 ng of NGF per g wet weight, respectively, and the densely innervated submandibular gland, heart atrium, and iris contained 0.5 +/- 0.1, 1.0 +/- 0.1, and 1.9 +/- 0.3 ng of NGF per g wet weight, respectively. Heart ventricle and skeletal muscle, which are poorly innervated by the sympathetic nervous system, did not contain detectable levels of NGF (less than 0.3 ng/g wet weight). Serum contained less than 0.05 ng of NGF per ml. The correlation between NGF levels and density of innervation is consistent with the concept that the production of NGF in target organs determines their density of innervation by the sympathetic nervous system.
本文描述了一种双位点酶免疫测定法，该方法不存在以往测定法中固有的假象，并且具有必要的高灵敏度，可用于测定交感神经系统及其靶器官中神经生长因子（NGF）的内源性水平。针对小鼠NGF（mNGF）的单克隆抗体和亲和纯化的多克隆抗体被共价连接到玻璃珠上作为第一个位点，并与β-半乳糖苷酶作为第二个位点偶联。通过检测荧光β-半乳糖苷酶反应产物，可以在每次测定中测定0.01 - 0.02 fmol的mNGF。添加到匀浆中的mNGF的回收率在50%至100%之间，具体取决于组织。发现大鼠颈上神经节和星状神经节每克湿重分别含有（平均值±标准误）25±4和19±3 ng的NGF，而神经密集分布的下颌下腺、心房和虹膜每克湿重分别含有0.5±0.1、1.0±0.1和1.9±0.3 ng的NGF。交感神经系统神经支配较少的心室和骨骼肌中未检测到可检测水平的NGF（低于0.3 ng/g湿重）。血清中每毫升含有的NGF低于0.05 ng。NGF水平与神经支配密度之间的相关性与靶器官中NGF的产生决定其由交感神经系统支配的密度这一概念一致。
