All lanes : Anti-Insulin Receptor beta antibody [C18C4] (ab69508) at 1/1000 dilution

Lane 2 : HeLa lysate (Human epithelial cell line from cervix adenocarcinoma)
Lane 3 : MCF-7 lysate (Human breast adenocarcinoma cell line)
Lane 4 : PC-12 lysate (Rat adrenal gland pheochromocytoma cell line)
Lane 5 : 3T3-L1 lysate (Mouse embryonic fibroblast cell line)

Developed using the ECL technique.

Predicted band size: 95 kDa
Observed band size: 98 kDa why is the actual band size different from the predicted?

This image was generated using the ascites version of the product.

IHC image of ab69508 staining in human normal kidney formalin fixed paraffin embedded tissue section, performed on a Leica Bond^TM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab69508, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

This image was generated using the ascites version of the product.

Overlay histogram showing HepG2 cells stained with ab69508 (red line). The cells were fixed with 80% methanol (5 min) and incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab69508, 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 IgG1 [ICIGG1] (ab91353, 2µg/1x10⁶ cells) used under the same conditions. Acquisition of >5,000 events was performed.

Please note that Abcam do not have any data for use of this antibody on non-fixed cells. We welcome any customer feedback.

This image was generated using the ascites version of the product.