Overlay histogram showing HepG2 cells stained with ab82591 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab82591, 1μg/1x106 cells) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-mouse IgG (H&L) (ab150113) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 1μg/1x106 cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.
ICC/IF image of ab82591 stained HepG2 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 (ab82591, 1/100 dilution) overnight at +4°C. The secondary antibody (green) was ab96879, DyLight® 488 goat anti-mouse 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.
Western blot - Grp75 antibody [30A5] (ab82591)
All lanes : Anti-Grp75 antibody [30A5] (ab82591) at 1/1000 dilution
Lane 1 : MWM Lane 2 : HeLa cell lysate (heat shocked) at 20 µg Lane 3 : L-929 cell lysate at 20 µg Lane 4 : Rat-2 cell lysate at 20 µg Lane 5 : CHO-K1 cell lysate at 20 µg Lane 6 : GPC-16 cell lysate at 20 µg
Predicted band size : 74 kDa Observed band size : 74 kDa
References for Anti-Grp75 antibody [30A5] (ab82591)
This product has been referenced in:
Lojewski X et al. Human iPSC models of neuronal ceroid lipofuscinosis capture distinct effects of TPP1 and CLN3 mutations on the endocytic pathway. Hum Mol Genet23:2005-22 (2014).
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