Recombinant Anti-CMT2 antibody [EPR9584] (ab150363)

False colour image of Western blot: Anti-CMT2 antibody [EPR9584] staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab150363 was shown to bind specifically to CMT2. A band was observed at 35 kDa in wild-type HeLa cell lysates with no signal observed at this size in MAD2L1BP knockout cell line ab265854 (knockout cell lysate ab257510). To generate this image, wild-type and MAD2L1BP knockout HeLa cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with primary antibodies overnight at 4°C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

ab150363 stained HeLa cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody ab150363 at 5µg/ml overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti- rabbit (ab96899) IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

Immunohistochemical analysis of paraffin embedded Human brain tissue labelling CMT2 binding protein with ab150363 antibody at a dilution of 1/100.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin embedded Human cervical carcinoma tissue labelling CMT2 binding protein with ab150363 antibody at a dilution of 1/100.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.