Westmark Cara J, Westmark Pamela R
Department of Neurology, University of Wisconsin, Madison, WI, USA.
Methods Mol Biol. 2019;1941:189-197. doi: 10.1007/978-1-4939-9077-1_13.
The use of synaptoneurosomes (SN) enables the detection of synaptic activity including the assessment of glutamate receptor function. SN are normally prepared by filtration and centrifugation methods. Here we review the preparation of SN by Percoll density gradient methodology for downstream applications that assesses glutamate receptor function such as measuring de novo protein synthesis. Major procedural steps include preparation of discontinuous Percoll-sucrose density gradients, collection of brain tissue, preparation of brain homogenates, isolation of synaptoneurosome bands from the discontinuous Percoll-sucrose gradients, and radiolabeling SN proteins. De novo protein synthesis can be reproducibly measured in SN prepared by this method.
使用突触神经小体（SN）能够检测突触活动，包括评估谷氨酸受体功能。SN通常通过过滤和离心方法制备。在此，我们回顾通过Percoll密度梯度方法制备SN的过程，用于评估谷氨酸受体功能的下游应用，如测量从头蛋白质合成。主要步骤包括制备不连续的Percoll-蔗糖密度梯度、收集脑组织、制备脑匀浆、从不连续的Percoll-蔗糖梯度中分离突触神经小体条带以及对SN蛋白进行放射性标记。通过这种方法制备的SN中，可以重复测量从头蛋白质合成。
