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The decrease in neuronal cell densities within the GCL was confirmed by neuronal staining using NeuN and TUJ1 antibodies.
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Typically, the purity of the neuronal cultures was greater than 95% as determined by neuronal staining with mouse anti-NeuN or goat anti-MAP2 (microtubule-associated protein 2) antibodies (Millipore); astrocytes were identified using a mouse anti-GFAP (glial fibrillary acidic protein) antibody (Sigma).
To identify neuronal cells expressing p65, immunofluorescence staining using anti-p65 plus anti-NeuN antibodies as a neuronal marker was performed for subjects with MCI-p.
Neuronal subpopulation expressing Dp71 was characterized by double-staining using presynaptic markers of mature excitatory (Type-1 vesicular glutamate transporter, VGLUT1) or inhibitory synapses GAD-65/67 GAD-65/67
The neuronal cells were stained using a immunofluorescence staining protocol established in Moore Lab, EPFL.
Neurons were selectively stained using antibodies directed against the neuronal protein Hu C/D.
(2) For visualization of neuronal cells, sections were stained using NeuroTrace green fluorescent Nissl stain (1 300; Molecular Probes, Netherlands; for 20 min) that binds to the Nissl substance, which is present exclusively in the somata of neuronal cells.
To assess neuronal death, sections were stained using FJB.
The neuronal soma was selected in the EGFP stain using the 'freehand selections' tool.
For histology, the sections were stained using a battery of techniques used for analysis of tissues in neuronal ceroid lipofuscinoses [ 4, 7, 8].
Histological sections were stained using hematoxylin and eosin stain (H&E stain) and observed by light microscopy.
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