碘标神经毒素—乙酰胆硷受体复合物对重症肌无力研究的初步应用

PRIMARY USE OF IODINE LABELED NEURO- TOXIN-ACETYLCHOLINE RECEPTOR COM- PLEX IN MYASTHENIA GRAVIS RESEARCH

眼镜蛇神经毒素化学性质稳定,易于保存,用^(191)I或^(125)I标记,方法简便,比活性高,能专一性地与菸硷型乙酰胆硷受体结合,常作为一种分子探测剂,用来研究蛇毒的毒理作用(杨钦照1977)、测定菸硷型乙酰胆硷受体的浓度,于临床还可以检测重症肌无力患者抗乙酰胆硷受体抗体(Lindstrom,1976;Lefvert 1978)。对研究神经肌接头的生理功能,阐明重症肌无力的发病机理、辅助诊断和观察疗效均有实际意义。

By means of chloramine T oxidation method, a-neurotoxin from Naja naja was labeled with 131I. After passing over a sephadex G-25 column and applied assending paper chromatograph, the rate of label reached to higher degree ( 80%) .The specific activity was 6μ.Ci/μ,g. With the help of two dimcntional double immunodiffusion and rocket immunoelectrophotesis self-illumination, its anljgcnicity still maintained was proved. Using the nonionic detergent and rate-zonal centrifuge, nicotinic acetylcholine receptor were purified from human skeletal muscle of the amputated extremity. The protein content of supernatant was 3.6mg/ml. This supernatant was incubated with various multiplied over-dose iodione labeled a-neurotoxin to prepare 131 I labeled to-xin-AChR complex.Passing over a sephadex G-200 column, its specific activity amounted to 110-378.5 pM/g.The iodine labeled neurotoxin and 5000 times unlabeled neurotoxin were incubated with AChR preparation resulting obvious competitive inhibi tion. Besides, there wasn’t any nonspecific binding between iodine labeled neurotoxin and bovine serum albumine. According to the method used in the anti-human globulin test, the 131l labeled neurotxin-AChR complex was incubated with the serum of mya-sthenia gravis. Then goast-anti-human globulin was added for immunoprecipitation. The antibodies binding to acetylcholine receptor (0.26 4 arbitrary units/1 ) were detected in the four of the five patients with generalized type of myasthenia gravis. This experiment demonstrated that iodine labeled neurotoxin used as to detect nico-tinic acetylcholine receptor and its antibody was characterized by its analysis up to 10 -12mole and definite specificity. It also proved that myasthenia gravis is as autoimmune disease. To detect nicotinic acetylcholine receptor antibody may become as an useful means in the diagnosis and also contributes 10 the investigation of therapy as an objective index.