Samples were digested with digestion solution in fume hood.
The tissue was digested using enzyme digestion solution containing 2.4 U/ml dispase, 650 U/ml type II collagenase and 10,000 Dornase U/ml DNase.
To test the absence of protease contamination in the PA28-αβ preparation, the substrate MBP102 129 was digested in the usual digestion solution without 20S proteasomes for 60 mins and no detectable degradation of the substrate was observed.
Fresh STs were minced and digested in tissue enzyme digestion solution as described previously [ 24].
The digestion solution was collected and digested for additional five times.
Calvaria were digested in 10 mL of digestion solution for 15 min at 37 °C with constant agitation.
After equilibration, each plug was digested at 25°C overnight in a digestion solution of 100 μl CutSmart buffer and 10 U of the restriction enzyme SmaI (NewEngland Biolabs, MA, USA).
The tissues were digested for 10 min with pepsin digestion solution (Dako) at room temperature, washed twice in distilled water for 5 min each, dehydrated in 70, 85 and 96% alcohol for 2 min each and dried.
Briefly, pellets were digested for 18 h at 60 °C, with a papain digestion solution containing 100 mM Sodium Phosphate Buffer supplemented with 5 mM Na2EDTA, 10 mM l-cysteine and 0.125 mg/ml papain (all from Sigma).
After removing the trypsin solution, the cartilage was digested at 37°C with type II collagenase using 1 ml of digestion solution per 100 mg tissue (Gibco, Grand Island, NY, USA).
After digestion solution was removed, tissue pieces were washed once with DMEM and digested with crude bacterial collagenase (type IA, C 2674; 1 mg/ml; Sigma, Saint Louis, MO USA).
