Cold is beneficial for healthy aging, finds new study UniCologne NatureAging
Worms were lysed in proteasome activity assay buffer using a Precellys 24 homogenizer . Human cells were collected in proteasome activity assay buffer and lysed by passing 10 times through a 27 G needle attached to a 1 ml syringe.for 10 min at 4 °C. For each sample, 25 μg of total protein were transferred to a 96-well microtiter plate and incubated with fluorogenic proteasome substrates. To measure trypsin-like proteasome activity, we used Ac-Arg-Leu-Arg-AMC .
Likewise, HEK293T cells were collected and lysed in non-denaturing lysis buffer supplemented with protease inhibitor cocktail. Cell lysates were homogenized by passing 10 times through a 27-gauge needle. Lysates from HEK293T cells expressing pARIS-mCherry-httQ23-GFP or pARIS-mCherry-httQ100-GFP were centrifuged at 8,000for 5 min at 4 °C. Lysates from HEK293T cells expressing FUS-HA-WT or FUS-HA-P525L were centrifuged at 1,000for 5 min at 4 °C.
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