Anti-Glutaminase C antibody [EPR19525] - BSA and Azide free

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Glutaminase C antibody [EPR19525] - BSA and Azide free (AB223129)

Immunofluorescent analysis of 100% methanol-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling Glutaminase C with ab202027 at 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic staining on HeLa cell line.

The nuclear counter stain is DAPI (blue). Tubulin is detected with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594)) at 1/200 dilution (red).

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) at 1/1000 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab202027).

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Glutaminase C antibody [EPR19525] - BSA and Azide free (AB223129)

Immunofluorescent analysis of 100% methanol-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling Glutaminase C with ab202027 at 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing mitochondrial staining on HeLa cell line.

The nuclear counter stain is DAPI (blue). COX IV is detected with ab33985 (anti-COX IV(mouse mAb)) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) at 1/1000 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab202027).

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Glutaminase C antibody [EPR19525] - BSA and Azide free (AB223129)

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab202027).

Immunofluorescent analysis of 4% PFA-fixed, 0.1% Triton X-100 permeabilized Hap1 WT and Hap1-GLS KO cells labelling Glutaminase C with ab202027 at 0.2 μg/ml concentration followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 (2 μg/ml dilution (Green). Image showing cytoplasmic staining in Hap1 WT cell line. ab7291 Anti-alpha Tubulin antibody [DM1A] was used to counterstain tubulin at 1/1000 dilution (Magenta). The nuclear counterstain was DAPI (Blue).

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-Glutaminase C antibody [EPR19525] - BSA and Azide free (AB223129)

This data was developed using ab202027, the same antibody clone in a different buffer formulation.

Flow cytometry overlay histogram showing left wild-type HAP1 positive cells and right negative GLS knockout HAP1 stained with ab202027 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilised with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS containing 10% normal goat serum to block non-specific protein-protein interaction followed by the antibody (ab202027) (1x 10⁶ in 100μl at 1 μg/ml (1/603)) for 30min at 22°C.

The secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed was incubated at 1/4000 for 30min at 22°C.

Isotype control antibody was Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control (black line) used at the same concentration and conditions as the primary antibody. Unlabelled sample (blue line) was also used as a control.

Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 530/30 bandpass filter.

This antibody gave a positive signal in wild-type HAP1 fixed with 4% formaldehyde (10 min) and then permeabilised with 0.1% PBS-Triton X-100 for 15 min under the same conditions.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-Glutaminase C antibody [EPR19525] - BSA and Azide free (AB223129)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling Glutaminase C with ab202027 at 1/600 dilution (red) compared with a rabbit monoclonal IgG isotype control (ab172730; black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody; blue). Goat anti rabbit IgG (Alexa Fluor^® 488) at 1/2000 dilution was used as the secondary antibody.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab202027).

• IP

Supplier Data

Immunoprecipitation - Anti-Glutaminase C antibody [EPR19525] - BSA and Azide free (AB223129)

Glutaminase was immunoprecipitated from 0.35 mg of HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate with ab202027 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab202027 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

Lane 1 : HeLa whole cell lysate, 10 μg (Input).

Lane 2 : ab202027 IP in HeLa whole cell lysate.

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab202027 in HeLa whole cell lysate.

Blocking and dilution buffer and concentration : 5% NFDM/TBST.

Exposure time : 10 seconds.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab202027).

All lanes:

Immunoprecipitation - Anti-Glutaminase C antibody [EPR19525] (<a href='/en-us/products/primary-antibodies/glutaminase-c-antibody-epr19525-ab202027'>ab202027</a>)

Observed band size: 65 kDa

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• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Glutaminase C antibody [EPR19525] - BSA and Azide free (AB223129)

Immunofluorescent analysis of 100% methanol-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 (Mouse embryonic fibroblast cell line) cells labeling Glutaminase C with ab202027 at 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic staining on NIH/3T3 cell line.

The nuclear counter stain is DAPI (blue). Tubulin is detected with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594)) at 1/200 dilution (red).

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) at 1/1000 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab202027).