Anti-Glutamine Synthetase 抗体 [3B6] - BSA and Azide free (ab64613)

IHC image of ab64613 staining in human tonsil 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 ab64613, 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.

Anti-Glutamine Synthetase antibody [3B6] - BSA and Azide free (ab64613) at 1 µg/ml + Jurkat cell lysate at 50 µg

Secondary
Goat Anti-Mouse IgG (H&L)-HRP at 1/2500 dilution

Predicted band size: 42 kDa
Observed band size: 37 kDa why is the actual band size different from the predicted?

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

ICC image of ab64613 stained PC12 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 (ab64613, 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.

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

Overlay histogram showing Jurkat cells stained with ab64613 (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 (ab64613, 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. This antibody gave a positive signal in Jurkat cells fixed with 4% paraformaldehyde (10 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

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

All lanes : Anti-Glutamine Synthetase antibody [3B6] - BSA and Azide free (ab64613)

Lane 1 : Glutamine Synthetase transfected lysate
Lane 2 : Non-transfected lysate

Predicted band size: 42 kDa

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

Anti-Glutamine Synthetase antibody [3B6] - BSA and Azide free (ab64613) + Raw 264.7 cell line

Predicted band size: 42 kDa

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

Anti-Glutamine Synthetase antibody [3B6] - BSA and Azide free (ab64613) + HepG2 cell lysate

Predicted band size: 42 kDa

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

Anti-Glutamine Synthetase antibody [3B6] - BSA and Azide free (ab64613) + PC-12 cell lysate

Predicted band size: 42 kDa

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

All lanes : Anti-Glutamine Synthetase antibody [3B6] - BSA and Azide free (ab64613) at 0.5 µg/ml

Lane 1 : Marker
Lane 2 : Zebrafish brain homogenate at 20 µg
Lane 3 : Zebrafish liver homogenate at 20 µg
Lane 4 : Zebrafish skeletal muscle homogenate at 20 µg
Lane 5 : JURKAT (Human T cell lymphoblast-like cell line) whole cell lysate at 20 µg

Secondary
All lanes : Goat polyclonal to Mouse IgG – H&L – Pre-Adsorbed (HRP) at 1/6000 dilution

Developed using the ECL technique.

Performed under reducing conditions.

Predicted band size: 42 kDa
Observed band size: 42 kDa


Exposure time: 4 minutes

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

Detection limit for recombinant GST tagged Glutamine Synthetase is approximately 0.03ng/ml as a capture antibody.

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