Anti kca3.1 p4997

To obtain cell lysates, cells were washed with ice‐cold PBS and incubated and harvested with a lysis buffer (150 mmol/L NaCl, 10 mmol/L NaH₂PO₄, 1 mmol/L EDTA, 1% TritonX100, 0.5% SDS) with protease inhibitor cocktail and phosphatase inhibitor (Sigma). Mouse and human brain were homogenized in RIPA buffer (ThermoFisher) with protease inhibitor cocktail and phosphatase inhibitor (Sigma) and used for Western blotting as described.9 The following primary antibodies (dilutions) were used: anti‐KCa3.1 P4997 (1:1000, Sigma), anti‐phospho P38MAPK and anti‐P38MAPK (1:1000, Cell Signaling), anti‐β actin (1:3000, Sigma), and anti‐GAPDH (1:2000, Cell Signaling). Secondary antibodies were HRP‐conjugated anti‐rabbit or anti‐mouse antibody (1:1000, GE Healthcare). Brain tissue samples from 5xFAD treated with senicapoc or control diet were fractionated into TBS‐soluble and TBS‐insoluble, SDS‐soluble fractions, which were used for Aβ42 quantification by a Human Aβ42 ELISA kit (Wako) as described.16 Alternatively, Aβ in each fraction was separated in 16.5% Tris/Tricine SDS gel electrophoresis (Bio‐Rad), transferred to PVDF membrane, and detected by anti‐Aβ (6E10, 1:1000, BioLegend).