Protease inhibitor cocktail tablets edta free

Human primary medulloblastoma cell lines, Daoy and UW228, were kindly donated by Dr Peter Dallas (Telethon Institute for Child Health Research, Subiaco, WA, Australia). RPMI 1640 and DMEM media, human recombinant insulin, glutamine, fetal bovine serums (FBS), molecular weight protein standards and NuPage 4%–12% Bis-Tris Mini gels were purchased from Invitrogen (Carlsbad, CA, USA). Protease inhibitor cocktail tablets (EDTA-free) were purchased from Roche Diagnostics (GmbH, Mannheim, Germany). Bradford dye reagent, Nonidet P40 (NP-40), 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) were purchased from Sigma-Aldrich (St. Louis, MO, USA).

The following primary antibodies were obtained from Santa Cruz Biotechnology Inc., USA; GR(H-300): sc-8992, PR(C-20) (which detects PRA and B isoforms): sc-539, AR(441): sc-7305, GAPDH(0411): sc-47724, MR(MCR, H300): sc-11412, ERα(MC-20): sc-542. The flotillin-1 (610820) antibody was purchased from BD Transduction Laboratories (USA). The following secondary antibodies were obtained from Santa Cruz Biotechnology Inc., USA; anti-mouse: sc-2005, anti-goat: sc-2350 (used as IgG for the ChIP assay) and anti-rabbit: sc-2313. The ligands dexamethasone (DEX), MPA, P4, NET-A, NET, aldosterone (ALD) and mibolerone (MIB) were obtained from Sigma-Aldrich (South Africa). Human tumour necrosis factor α (TNFα) was obtained from Celtic Diagnostics (South Africa). Protease inhibitor cocktail tablets (EDTA-free) (cat #04693159001) were obtained from Roche (South Africa). Cycloheximide (CHX) was purchased from Sigma-Aldrich (South Africa).