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Membranes were then incubated with a primary antibody (Rabbit anti-GST (Cell Signaling Technology 91G1) diluted 1:2500, or Mouse anti-FLAG (Sigma F1804) diluted 1:1000 in blocking buffer for 1 h, washed 3x with 25 mL TBS-T, then incubated with a secondary antibody (Anti-Rabbit HRP-linked IgG (Cell Signaling Technology 7074S) diluted 1:2000, or Anti-Mouse HRP-linked IgG (Invitrogen 62-6520) diluted 1:2000) for 1 hr, and washed 3x again with 25 mL TBS-T per wash

HT affects more than 15 % of females over 60 years and 2 % of males ( Reference Nacamulli, Petricca and Mian 6 )

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