Anti-67kDa Laminin Receptor antibody [MLuC5] (ab3099)

Immunocytochemistry/ Immunofluorescence - 67kDa Laminin Receptor antibody [MLuC5] (ab3099)

ICC/IF image of ab3099 stained MCF7 cells. The cells were 100% methanol fixed (5 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 (ab3099, 5µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-mouse IgG (H+L) used at a 1/1000 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 - 67kDa Laminin Receptor antibody [MLuC5] (ab3099)

Anti-67kDa Laminin Receptor antibody [MLuC5] (ab3099) at 1/1000 dilution + Mouse pancreatic cancer tissue lysate at 35 µg

Secondary
HRP-conjugated Sheep anti-Mouse polyclonal at 1/2000 dilution
developed using the ECL technique

Performed under reducing conditions.

Predicted band size : 33 kDa
Observed band size : 30 kDa (why is the actual band size different from the predicted?)


Exposure time : 10 minutes

This image is courtesy of an anonymous Abreview.

Flow Cytometry - 67kDa Laminin Receptor antibody [MLuC5] (ab3099)

Overlay histogram showing MCF7 cells stained with ab3099 (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 (ab3099, 1µg/1x10⁶ cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgM [ICIGM] (ab91545, 2µg/1x10⁶ cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in MCF7 cells fixed with 80% methanol (5 min)/permeabilized in 0.1% PBS-Tween used under the same conditions.