Characterization of Mitochondrial and Cytoskeletal Proteins Using Antibodies

The following polyclonal rabbit antisera directed against the indicated antigens were produced in our lab and have been used before [23 (link), 35 (link)]. The working dilutions for immunoblots (IB) and IF are indicated: VDAC (IB 1:1,000), ATOM40 (IB 1:10,000; IF 1:1,000), cytochrome C (IB 1:1,000), ATOM69 (IB 1:50), LipDH (IB 1:10,000) and CoxIV (IB 1:1,000). Commercially available monoclonal antibodies were used as follows: mouse c-Myc (Invitrogen, 132500; IB 1:2,000; IF 1:50), mouse HA (Enzo Life Sciences AG, CO-MMS-101 R-1000; IB 1:5,000; IF 1:1,000) and mouse EF1a (Merck Millipore, Product No. 05–235; IB 1:10,000). Monoclonal anti-tyrosinated α-tubulin antibody YL1/2 [54 (link)] (IFA 1:500) produced in rat was a generous gift from Prof. Keith Gull, University of Oxford.
Secondary antibodies for IB analysis were IRDye 680LT goat anti-mouse, IRDye 800CW goat anti-rabbit (LI-COR Biosciences, 1:20,000) and horse radish peroxidase-coupled goat anti-mouse and anti-rabbit (Sigma-Aldrich, 1:5,000). Secondary antibodies for IF were goat anti-mouse Alexa Fluor 633, goat anti-mouse Alexa Fluor 596, goat anti-rabbit Alexa Fluor 488 and goat anti-rat Alexa Fluor 488 (all from ThermoFisher Scientific, 1:1000)