Lenses were dissected in PBS, and homogenized in PBS, 4 mM EDTA, 2 mM phenylmethanesulfonyl fluoride, and cOmplete EDTA-free protease inhibitor cocktail (Roche Applied Science, Indianapolis, IN) in a glass-glass homogenizer, and then sonicated. Homogenates were stored at −80°C until use. Protein concentrations were determined using the BioRad Protein Assay (BioRad, Hercules, CA) based on the Bradford dye-binding procedure.
20 Immunoblots were performed as described in Minogue et al.
21 using anti-Cx50, anti-Cx46, anti-AQP0, anti-H3 histone, anti-PDI, anti-Tom20, or anti-crystallin antibodies. Except as noted, immunoblots were performed in triplicate, quantified by densitometry, and are reported in the text as percentages of the values determined from wild-type samples.