The localization of proteins by immunofluorescence in fixed HOS 143B cells was performed as described previously (Minczuk, 2010 (link)). The following antibodies were used: rabbit anti-TOM20 (Santa Cruz), Alexa Fluor 488 anti-rabbit mouse anti-FLAG (Sigma, F1804, 1:200), Alexa Fluor 594 anti-mouse (Life Technologies, A11006, 1:200), rat anti-HA (Roche, 11867431001, 1:200), Alexa Fluor 594 anti-rat (Life Technologies, A11001, 1:200). Immunofluorescence images were captured using a Nikon N-SIM confocal microscope.
Western blotting analysis was performed as previously (Minczuk et al, 2011 (link)), and membranes were probed with the following primary antibodies: anti-FLAG and anti-HA as above, mouse anti-OXPHOS cocktail (Mitosciences, MS601, 1:300), mouse anti-TOM22 (Abcam, ab10436, 1:5000), mouse anti-β-actin (Sigma, A2228, 1:100,000), rabbit anti-SSB1 (kindly donated by Prof. D. Kang, 1:4000), rabbit anti-histone H4 (Abcam, ab10158; 1:5000). Secondary antibodies used were HRP-conjugated goat antibodies to rabbit (Promega, W401B; 1:2000), mouse (Promega, W402B, 1:2000) and rat (Santa Cruz, SC2065, 1:1000).
Western blotting analysis was performed as previously (Minczuk et al, 2011 (link)), and membranes were probed with the following primary antibodies: anti-FLAG and anti-HA as above, mouse anti-OXPHOS cocktail (Mitosciences, MS601, 1:300), mouse anti-TOM22 (Abcam, ab10436, 1:5000), mouse anti-β-actin (Sigma, A2228, 1:100,000), rabbit anti-SSB1 (kindly donated by Prof. D. Kang, 1:4000), rabbit anti-histone H4 (Abcam, ab10158; 1:5000). Secondary antibodies used were HRP-conjugated goat antibodies to rabbit (Promega, W401B; 1:2000), mouse (Promega, W402B, 1:2000) and rat (Santa Cruz, SC2065, 1:1000).
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