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After incubation on ice for 15 min, 25 l of 10% (v/v) Nonidet P-40 (Nacalai Tesque, Kyoto, Japan) was added to the cell suspension, followed by vortexing for 10 s at maximum speed

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High alcohol intake can also cause your voice to age prematurely

The first song, The Bitter End, is kind of autobiographical, Carter said