To ensure full equilibration of minerals and the specific chelator, pre-washing was maintained in EDTA-2Na·2H2O for 2.5 h with agitation during the first 30 min [42], in 2,2'-bipyridyl for 2 h with continuous agitation, and in deionized H2O for 48 h without agitation.
Since then the heat from Earth's mantle has kept the plates in a state of continuous agitation.
Gel destaining was performed by continuous agitation in a methanol: acetic acid: water (20 10 70 v/v/v) solvent until obvious bands of proteins were obtained.
Kinase assays were performed for 30 minutes at 30°C under continuous agitation in kinase buffer containing 200 µM ATP and 1 µg of GSK-3 fusion protein.
Bacterial stocks were prepared by growing strains at 37°C with continuous agitation in Middlebrook 7H9 broth supplemented with 10% OADC, 0.2% glycerol and 0.05% Tween 80 or further supplemented with 80 µg/ml lysine and 25 µg/ml pantothenate (for mc26020).
The method involved slow addition of polymer in cold water with continuous agitation.
The exacerbation of schizophrenia with acute psychotic agitation was added as an inclusion criterion, and continuous agitation in patients for at least 2 weeks duration before informed consent was to be signed was added as an exclusion criterion.
Kinase assays were performed for 30 min at 30°C under continuous agitation in kinase buffer containing 200 μM ATP and 1 μg of GSK-3 fusion protein according to the manufacturer's instructions (Cell Signaling Technology, Beverly, MA).
The broth was fermented at 32 °C for 72 h in an automatic incubator with continuous agitation (150 rpm).
The slurry was incubated at 32 °C for 72 h in an automatic incubator with continuous agitation (150 rpm), and 2 mL of samples were drawn at 4, 8, 12, 24, 48, and 72 h to monitor the fermentation.
