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Spinal section were then incubated with a mixture of Alexa Fluor 488-labeled chicken anti-mouse IgG antibody (1 400, cat. A-21200; Molecular Probes, Eugene, OR, USA; green fluorescence) and DyLight 549-conjugated donkey anti-rabbit IgG antibody (1 400, cat. 711-506-152; Jackson ImmunoResearch Laboratories Inc., West Grove, PA, USA; red fluorescence) for 40 min at room temperature.

The number of ß-amyloid-positive structures present in each section were then quantitated using color selection to separate the plaques from background labeling.

Formalin-fixed sections (human section) were, then, treated with target retrieval solution (DAKO, Carpinteria, CA) for 10 min in steamer; no target retrieval treatment for zinc-based Beckstead's solution-fixed mouse tissues.

Section were then washed in phosphate buffered saline and incubated for 2 hours at room temperature with the biotinylated anti-rabbit secondary antibody (Sigma-Aldrich, St Louis, MO, USA) diluted 1∶250 in phosphate buffered saline.

All 200× images that composed an entire lymph node section were then analyzed using a custom image analysis software, GemIdent, yielding the total number of cells for each stained phenotype and the location (x, y coordinate) of each cell identified [5].

Average values across groups within each section were then calculated.

The section was then counterstained with haematoxylin.

The section was then examined using a fluorescence microscope.

Sections were then counterstained with hematoxylin.

Sections were then counterstained by hematoxylin.

Sections were then imaged with TissueFax.