一种用于检测人类大脑中苯乙醇胺N-甲基转移酶的高灵敏度检测方法。
A highly sensitive assay for phenylethanolamine N-methyltransferase in human brain.
作者信息
Burke W J, Hanson D M, Chung H D
出版信息
Proc Soc Exp Biol Med. 1986 Jan;181(1):66-70. doi: 10.3181/00379727-181-42225.
A rapid, highly sensitive assay for phenylethanolamine N-methyltransferase in brain using the natural substrate, norepinephrine, is described. The method is based on the selective adsorption and elution of the reaction product, epinephrine, from alumina. A small but important further lowering of blanks and increase in sensitivity is attained by removal of the radiolabeled substrate, [methyl-3H]-S-adenosylmethionine by precipitation as the reineckate prior to adsorption of norepinephrine to alumina. The assay has a sensitivity of 30 fmole and the PNMT activity could be measured in as little as 1 mg (wet wt) of human locus coeruleus tissue. The sensitivity is enhanced by homogenizing tissue in small volumes and removing potential inhibitors by dialysis. We report for the first time PNMT activity in specific regions of the human cerebral and cerebellar cortex.
本文描述了一种使用天然底物去甲肾上腺素对脑中苯乙醇胺N-甲基转移酶进行快速、高度灵敏的检测方法。该方法基于从氧化铝上选择性吸附和洗脱反应产物肾上腺素。在去甲肾上腺素吸附到氧化铝之前，通过以雷氏盐沉淀的方式去除放射性标记底物[甲基-³H]-S-腺苷甲硫氨酸，可进一步小幅但显著地降低空白值并提高灵敏度。该检测方法的灵敏度为30飞摩尔，在低至1毫克（湿重）的人蓝斑组织中即可检测到苯乙醇胺N-甲基转移酶活性。通过小体积匀浆组织并通过透析去除潜在抑制剂可提高灵敏度。我们首次报道了人大脑和小脑皮质特定区域的苯乙醇胺N-甲基转移酶活性。
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