Tetramethylbenzidine is used as peroxidase substrate. Finally, an acidic stop solution is added to terminate the reaction. The colour changes from blue to yellow. The intensity click here of the yellow colour is directly proportional to the concentration of α1-antitrypsin. Samples are quantified by referring their optical density to a lot-dependant master calibration curve and the use of a calibrator that is run with each test. Data are expressed in mg/dL. Analyses of blood parameters CP was analyzed with a

commercially available ELISA (Immundiagnostik AG, Bensheim, Germany) via reaction of protein with dinitrophenylhydrazine (DNPH). The non-protein constituents and unconjugated DNPH are separated by ultracentrifugation. The proteins are adsorbed to an ELISA plate and incubated with anti-DNPH antibody followed by antibody-linked horseradish peroxidase. Absorbances are related to a standard curve prepared with oxidized serum albumin. The carbonyl protein content is calculated from the estimated carbonyl concentration and the total protein content of the sample. For this reason, a parallel determination of the protein content is required. Data are expressed in pmol/mg. MDA was determined according to a previously described HPLC method by Pilz et al. [29] after derivatization with 2,4-DNPH. This method determines the protein bound MDA. The HPLC separations were performed

with an L-2200 autosampler, a L-2130 HTA pump and a L-2450 diode array detector (all: VWR Hitachi Vienna; Austria). Lepirudin Detector signals (absorbance at 310 nm) were recorded and program EZchrom Elite (VWR) was used for data requisition and analysis. Data are expressed in nmol/mL. selleck kinase inhibitor Analysis of TOS: This assay (Immundiagnostik AG, Bensheim, Germany) determines total lipid peroxides and is performed by the reaction of a peroxidase with the peroxides in the sample followed by the conversion of tetramethylbenzidine to a colored product. After addition of a stop solution the samples are measured at

450 nm in a microtiter plate reader. The quantification is performed by the delivered calibrator. Data are expressed in μmol/L H2O2. TNF-α was analyzed with a commercially available ELISA (Immundiagnostik AG, Bensheim, Germany) allowed quantitative determination of Tumor Necrosis Factor-α by using monoclonal antibodies and a horseradish peroxidase labeled conjugate. The amount of the converted substrate by the peroxidase is directly proportional to the amount of bound TNF-α and can be determined photometrically. Data are expressed in pg/mL. IL-6 was also measured with commercial available ELISA kits (Invitrogen, LifeTech Austria, Vienna, Austria) using monoclonal antibodies specific for human IL-6. Based on the binding of streptavidin-peroxidase to antibodies the intensity of a colored see more adduct is directly proportional to the concentration of the cytokine and can be determined photometrically. Data are expressed in pg/mL.