Cell based proteasome glo chymotrypsin like kit

Eight 6-week-old athymic nude mice were intraperitoneally injected with either 200 µg/kg MRZ or an equivalent volume of vehicle (2% Dimethyl sulfoxide [DMSO] in saline) and sacrificed 1 hour later. The cerebellum of each mouse was extracted, mechanically disaggregated, and passed through a 70-µM cell filter. Cells were then aliquoted at 10k cell/well of a 96-well plate and assayed using the Cell-Based Proteasome-Glo Chymotrypsin-like kit (Promega).

Cells were seeded in 96-well plates, treated with 100 nM MRZ and/or 10 mM NaC for either 6 or 24 hours, and then proteasome activity assayed using the Cell-Based Proteasome-Glo Chymotrypsin-like kit (Promega).

To measure proteasome activity, the Proteasome-Glo chymotrypsin-like cell-based kit (Promega Biotech Ibérica, Madrid, Spain) was used according to the manufacturer’s instructions. HEK293T cells were seeded in a white 96-well plate, and after transfection, they were treated with 4QMn at 20 µM for 24 h. After the incubation period, cells were processed according to the protocol of the kit. When the activity of the proteasome wanted to be inhibited with a possible rescued capacity of 4QMn, the proteasome inhibitor, MG132 (M7449, Sigma Aldrich, Darmstadt, Germany) was used at 10 µM. In this case, cells were treated with 4QMn and MG132 simultaneously.