Human mouse tgf β1 elisa ready set go kit

Activation of TGF-β in cell culture experiments or in BALF obtained from WT and Del-1^−/− mice with PF induced by inactive TGF-β-expressing adenovirus was measured using the human/mouse TGF-β1 ELISA Ready-SET Go kit (eBioscience) according to the manufacturer's protocol. Briefly, the plate was coated overnight at 4°C with capture antibody, washed five times with PBST (0.05% Tween 20 in PBS), blocked with 1× assay buffer, incubated at RT for 1 h, and washed five times with 0.05% PBST. The plate was then loaded with samples and standards, incubated at RT for 2 h or at 4°C overnight, washed five times with 0.05% PBST, and incubated at RT for 1 h with biotin-conjugated anti-active TGF-β antibody. After washing five times with 0.05% PBST, the plate was incubated at RT for 30 min with avidin-conjugated HRP. The plate was washed five times with 0.05% PBST, developed with TMB, and read at 630 nm on a microplate reader (Synergy HT). The concentrations of LAP were measured as described above but using a mouse LAP (TGF beta 1) Ready-SET Go kit (eBioscience).

Pieces of pancreas were removed, weighed and placed into a tube containing Complete Protease Inhibitor Cocktail (Roche Diagnostics, Abbott Park, IL, USA). Pancreatic tissue was homogenized using a Polytron homogenizer (Kinematica, Luzern, Switzerland) and interleukin (IL)-2, IL-6, interferon gamma, IL-17, IL-4 and IL-10 levels were detected in the supernatant by the cytometric bead array method (Th1/Th2/Th17 kit; BD), according to the manufacturer’s instructions. The concentration of transforming growth factor beta (TGF-β) in pancreatic tissue was determined using the Human/Mouse TGF-β1 ELISA Ready-Set-Go kit (eBioscience, San Diego, CA, USA). Serum cytokine levels were also determined by the cytometric bead array method.