Identification and Characterization of Amyloid-β Accumulation in Synaptic Mitochondria

Mitochondrial and synaptic dysfunction is an early pathological feature of Alzheimer's disease (AD). Accumulation of amyloid beta-peptide (Aβ) in mitochondria, particularly in synaptic mitochondria, potentiates and amplifies synaptic injury and disruption of synaptic transmission, leading to synaptic dysfunction and ultimately to synaptic failure. Thus, determination of the presence and levels of Aβ in synaptic mitochondria associated with amyloid pathology is important for studying mitochondrial amyloid pathology. Here, we present a detailed methodology for the isolation of synaptic mitochondria from brain tissues and the determination of Aβ levels in the isolated mitochondria as well as ultrastructural localization of synaptic mitochondrial Aβ. These methods have been used successfully for the identification and characterization of Aβ accumulation in synaptic mitochondria from mouse brains derived from transgenic AD mouse model. Additionally, we comprehensively discuss the sample preparation, experimental details, our unique procedures, optimization of parameters, and troubleshooting.