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. Author manuscript; available in PMC: 2008 Aug 1.

Published in final edited form as: Exp Eye Res. 2007 May 21;85(2):270–279. doi: 10.1016/j.exer.2007.05.001

Fresh pig lenses were incubated in TC199 medium with or without 0.2 mM H₂O₂ for 24hours. The concentration of H₂O₂ was maintained using 2.31 unit of glucose oxidase in the medium. The lenses were taken at the indicated time points and the epithelial layers were removed. A, Effect of H₂O₂ on protein expression. The total soluble protein fraction was prepared and 40μg of protein from each time point was subjected to immunoblot analysis with antibodies specific to TRx and TBP-2. Glyceraldehyde-3-phosphate dehydrogenase (G3PD) was used as an internal control. B, Effect of H₂O₂ on mRNA expression. Lenses were removed at the indicated times and epithelial layers separated. Total RNA was extracted from the three pooled epithelial layers (in each group) and reverse transcribed. Primers were designed and synthesized to detect TBP-2, TRx and β-actin. Equal amounts of synthesized cDNA were amplified by PCR with synthesized primers. PCR products were analyzed by agarose gel electrophoresis. β-actin was amplified as an internal control.

Fresh pig lenses were incubated in TC199 medium with or without 0.2 mM H₂O₂ for 24hours. The concentration of H₂O₂ was maintained using 2.31 unit of glucose oxidase in the medium. The lenses were taken at the indicated time points and the epithelial layers were removed. A, Effect of H₂O₂ on protein expression. The total soluble protein fraction was prepared and 40μg of protein from each time point was subjected to immunoblot analysis with antibodies specific to TRx and TBP-2. Glyceraldehyde-3-phosphate dehydrogenase (G3PD) was used as an internal control. B, Effect of H₂O₂ on mRNA expression. Lenses were removed at the indicated times and epithelial layers separated. Total RNA was extracted from the three pooled epithelial layers (in each group) and reverse transcribed. Primers were designed and synthesized to detect TBP-2, TRx and β-actin. Equal amounts of synthesized cDNA were amplified by PCR with synthesized primers. PCR products were analyzed by agarose gel electrophoresis. β-actin was amplified as an internal control.