Correlative Light and Electron Microscopy to Study Microglial Interactions with β-Amyloid Plaques.
A detailed protocol is provided here to identify amyloid Aβ plaques in brain sections from Alzheimer's disease mouse models before pre-embedding immunostaining (specifically for ionized calcium-binding adapter molecule 1 (IBA1), a calcium binding protein expressed by microglia) and tissue processing for electron microscopy (EM). Methoxy-X04 is a fluorescent dye that crosses the blood-brain barrier and selectively binds to β-pleated sheets found in dense core Aβ plaques. Injection of the animals with methoxy-X04 prior to sacrifice and brain fixation allows pre-screening and selection of the plaque-containing brain sections for further processing with time-consuming manipulations. This is particularly helpful when studying early AD pathology within specific brain regions or layers that may contain very few plaques, present in only a small fraction of the sections. Post-mortem processing of tissue sections with Congo Red, Thioflavin S, and Thioflavin T (or even with methoxy-X04) can label β-pleated sheets, but requires extensive clearing with ethanol to remove excess dye and these procedures are incompatible with ultrastructural preservation. It would also be inefficient to perform labeling for Aβ (and other cellular markers such as IBA1) on all brain sections from the regions of interest, only to yield a small fraction containing Aβ plaques at the right location. Importantly, Aβ plaques are still visible after tissue processing for EM, allowing for a precise identification of the areas (generally down to a few square millimeters) to examine with the electron microscope.
Neurosciences Axis, CHU de Québec Research Center.
Kanchan
K
Bisht
Neurosciences Axis, CHU de Québec Research Center.
Hassan
H
El Hajj
Neurosciences Axis, CHU de Québec Research Center.
Julie C
JC
Savage
Neurosciences Axis, CHU de Québec Research Center.
Maria G
MG
Sánchez
Neurosciences Axis, CHU de Québec Research Center; Tremblay.Marie-Eve@crchudequebec.ulaval.ca.
Marie-Ève
MÈ
Tremblay
D000544:Alzheimer Disease / Q000175:diagnosis; D016229:Amyloid beta-Peptides; D000818:Animals; D001812:Blood-Brain Barrier; D001921:Brain; D051379:Mice; D017628:Microglia; D008854:Microscopy, Electron; D058225:Plaque, Amyloid
D016229:Amyloid beta-Peptides