Tnf α elisa kit

Bronchial epithelial cell growth medium (BEGM) with antibiotics was purchased from Lonza (Walkersville, MD). The BEGM was prepared following manufacturer’s guideline, which contained all the supplements (BPE, hEGF, epinephrine, transferrin, insulin, retinoic acid, triiodothyronine, GA) except hydrocortisone to avoid any inhibitory effect of corticosteroids on cell pro-inflammatory responses. RNA lysis buffer RLT was from Qiagen (Hilden, Germany). RIPA western lysis buffer was purchased from Thermo-Fisher Scientific (Waltham, MA). DMEM (high glucose) for making D10 (DMEM + 10% FBS + 1% Pen/Strep + 1% Amphotericin B + 1% L-Glutamine+ 0.5% Gentamicin) was from GE Life Sciences (Logan, UT). The nuclear extraction kit and TransAM NF-κB p65 assay kit were from Active Motif (Carlsbad, CA). IL-8, IP-10 and TNF-α ELISA kits were obtained from R&D systems (Minnieaplois, MN).

Cytokine concentrations from the harvested supernatants of RASFs after appropriate treatment were measured by using commercial interleukins IL-6, IL-1β and IL-8 and tumor necrosis factor-α (TNF-α) ELISA kits (R&D Systems, Minneapolis, USA) according to the instructions of the manufacturer. Each sample was prepared in triplicate, and the experiment was repeated three times.

Blood was collected retro-orbitally under anesthesia immediately prior to sacrifice. Serum was obtained using serum separator tubes with lithium heparin (Becton Dickinson) and kept at -80C. ELISA kits were purchased from: Mouse adiponectin, leptin, adipsin, resistin and TNFα ELISA kits from R&D; Mouse Viafatin and Insulin ELISA kit from Ray biotech (Norcross, GA); Serum CTx-1 and TRAP5b kits from Nordic Bioscience; osteocalcin ELISA kit from Biomedical Technologies Inc.