Systemic immune challenges trigger and drive Alzheimer-like neuropathology in mice

Table 3 Western blotting parameters

Antibody	Run time, hours/voltage	Transfer time hours	Blocking reagent/time, hours	Secondary antibody incubation, hours
total Tau/Ab-2	1.5 to 2/125	1.5	10% WBR¹/1	1.5
PHF-Tau
pTau^T205
APP, C-terminal	2/125 or 2.5/100³	1.5	10% FBS²/2	1
APP (A4)	3/1253	1.5	10% WBR/1	1.5
β-Actin⁴			10% WBR/1	1

FBS, fetal bovine serum (heat-inactivated); paired helical filament; WBR, Western blocking reagent (Roche Applied Science).
¹Incubation of membrane in boiling PBS for 5 minutes recommended.
²Incubation of membrane in boiling PBS for 5 minutes required.
³Western blotting apparatus kept in the water bath at room temperature to prevent overheating.
⁴β-actin was measured for each blot after stripping (2% SDS at 56°C for 30 minutes).

ISSN: 1742-2094