Anti-PGP9.5 antibody [BH7] (ab72911)

Overlay histogram showing SH-SY5Y cells stained with ab72911 (red line). The cells were fixed with 4% paraformaldehyde (10 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 (ab72911, 1μg/1x10⁶ cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H&L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2μg/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. This antibody gave a positive signal in SH-SY5Y cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

Anti-PGP9.5 antibody [BH7] (ab72911) at 1/5000 dilution + rat hippocampal homogenate

Predicted band size: 25 kDa
Observed band size: <43 kDa why is the actual band size different from the predicted?

ab72911 at 1/500 dilution staining PGP9.5 in rat spinal cord by Immunofluorescence (shown in red). Anti-neurofilament NF-H antibody staining is shown in green. The large cells are a-motorneurons and PGP9.5 fills the cytoplasm of their perikarya and dendrites.