کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
1216195 | 1494182 | 2007 | 10 صفحه PDF | دانلود رایگان |
We noted differences in the antibody response to 3-nitrotyrosine (NO2Tyr) in fixed and non-fixed tissues, and studied therefore potential problems associated with non-fixed tissues in Western blot analyses. Three different monoclonal anti-nitrotyrosine antibodies in Western blot analysis of inflammatory stimulated rat abdominal, liver and lung tissue homogenates caused no immunoreactivity, in contrast to a polyclonal nitrotyrosine antibody applied in fixed and non-fixed tissues. Western blot studies using both mono- and polyclonal antibodies showed a temperature- and heme group-dependent reduction of NO2Tyr in nitrated rat and bovine serum albumin incubated with dithiothreitol. Mass spectrometric analyses of a nitrated peptide angiotensin II revealed under similar conditions a positive temperature effect between 56 and 70 °C on reduction of NO2Tyr to 3-aminotyrosine which is not detected by anti-NO2Tyr antibodies. Western blot analysis may therefore underestimate the level of tissue nitration, and factors causing a reduction of NO2Tyr during sample preparation might conceal the actual nitration of proteins.
Journal: Journal of Chromatography B - Volume 851, Issues 1–2, 15 May 2007, Pages 277–286