All lanes : Anti-ABAT/GABA-T antibody [EPR20842] (ab216465) at 1/1000 dilution

Lane 1 : Mouse liver lysate
Lane 2 : Rat liver lysate
Lane 3 : Rat brain lysate
Lane 4 : Human fetal liver lysate

Lysates/proteins at 10 µg per lane.

Secondary
Lanes 1-2 : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Lane 3 : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Lane 4 : VeriBlot for IP Detection Reagent (HRP) (ab131366) at 1/1000 dilution

Predicted band size: 56 kDa

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

Exposure time: Lanes 1 and 2: 3 seconds; Lane 3: 5 seconds; Lane 4: 26 seconds.

The observed molecular mass is consistent with what has been described in the literature (PMID: 11459221).

Immunofluorescent analysis of 100% methanol-fixed HepG2 (human hepatocellular carcinoma epithelial cell) cells labeling ABAT/GABA-T with ab216465 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 (green). Confocal image showing mitochondrial staining in HepG2 cell line is observed. The nuclear counter stain is DAPI (blue).

Mitochondria are stained with ab33985 Anti-COX IV (mouse mAb) - Mitochondrial Marker followed by ab150120 AlexaFluor^®594 Goat anti-Mouse secondary both at 1/1000 dilution (red).

-ve control 1: ab216465 at 1/100 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.

-ve control 2: ab33985 Anti-COX IV (mouse mAb) - Mitochondrial Marker at 1/1000 dilution followed by ab150077 (AlexaFluor®488 Goat anti-Rabbit secondary) at 1/1000 dilution.

Immunofluorescent analysis of 100% methanol-fixed MCF7 (human breast adenocarcinoma epithelial cell) cells labeling ABAT/GABA-T with ab216465 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 (green). Confocal image showing mitochondrial staining in MCF7 cell line. The nuclear counter stain is DAPI (blue).

Mitochondria are stained with ab33985 Anti-COX IV (mouse mAb) - Mitochondrial Marker followed by ab150120 AlexaFluor^®594 Goat anti-Mouse secondary both at 1/1000 dilution (red).

-ve control 1: ab216465 at 1/100 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.

-ve control 2: ab33985 Anti-COX IV (mouse mAb) - Mitochondrial Marker at 1/1000 dilution followed by ab150077 (AlexaFluor®488 Goat anti-Rabbit secondary) at 1/1000 dilution.

All lanes : Anti-ABAT/GABA-T antibody [EPR20842] (ab216465) at 1/1000 dilution

Lane 1 : Mouse brain lysate
Lane 2 : Human brain lysate
Lane 3 : MCF7 (human breast adenocarcinoma epithelial cell) whole cell lysate
Lane 4 : HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate
Lane 5 : SH-SY5Y (human neuroblastoma epithelial cell) whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 56 kDa

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

Exposure time: Lane 1,2,3 and 4: 6 seconds; Lane 5: 92 seconds.

The observed molecular mass is consistent with what has been described in the literature (PMID: 11459221).

ABAT/GABA-T was immunoprecipitated from 0.35 mg MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysate with ab216465 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab216465 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/5,000 dilution
Lane 1: MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysate 10 μg (Input).
Lane 2: ab216465 IP in MCF7 whole cell lysate (+).
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab216465 in MCF7 whole cell lysate (-).

Blocking/Dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 3 minutes.

 

Flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol-permeabilized MCF7 (Human breast adenocarcinoma epithelial cell) cells labeling ABAT/GABA-T with ab216465 at 1/600 (red) compared with a Rabbit monoclonal IgG (ab172730) (black) and an unlabelled control (cells incubated with secondary antibody only) (blue). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077), at 1/2000 dilution was used as the secondary antibody.

Flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol-permeabilized HepG2 (Human hepatocellular carcinoma epithelial cell) cell line labeling ABAT/GABA-T with ab216465 at 1/60 (red) compared with a Rabbit monoclonal IgG (ab172730) (black) and an unlabelled control (cells incubated with secondary antibody only) (blue). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077), at 1/2000 dilution was used as the secondary antibody.

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen mouse liver tissue labeling ABAT/GABA-T with ab216465 at 1/500 dilution (green), followed by ab150077 AlexaFluor^®488 Goat anti-Rabbit secondary at a 1/1000 dilution. Cytoplasmic staining in rat liver is observed. Counter stained with DAPI (blue).
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ab150077 AlexaFluor^®488 Goat anti-Rabbit used at a 1/1000 dilution.

Perform heat-mediated antigen retrieval by using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen rat choroid plexus tissue labeling ABAT/GABA-T with ab216465 at 1/500 dilution (green), followed by ab150077 AlexaFluor^®488 Goat anti-Rabbit secondary at a 1/1000 dilution. Cytoplasmic staining in rat choroid plexus (PMID:25239459, PMID: 11459221) is observed. Counter stained with DAPI (blue).
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ab150077 AlexaFluor^®488 Goat anti-Rabbit used at a 1/1000 dilution.

Perform heat-mediated antigen retrieval by using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen mouse choroid plexus tissue labeling ABAT/GABA-T with ab216465 at 1/500 dilution (green), followed by ab150077 AlexaFluor^®488 Goat anti-Rabbit secondary at a 1/1000 dilution. Cytoplasmic staining in mouse choroid plexus (PMID:25239459, PMID: 11459221) is observed. Counter stained with DAPI (blue).
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ab150077 AlexaFluor^®488 Goat anti-Rabbit used at a 1/1000 dilution.

Perform heat-mediated antigen retrieval by using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen mouse cerebellum tissue labeling ABAT/GABA-T with ab216465 at 1/500 dilution (green), followed by ab150077 AlexaFluor^®488 Goat anti-Rabbit secondary at a 1/1000 dilution. Cytoplasmic staining in mouse cerebellum (PMID:25239459, PMID: 11459221) is observed. Counter stained with DAPI (blue).
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ab150077 AlexaFluor^®488 Goat anti-Rabbit used at a 1/1000 dilution.

Perform heat-mediated antigen retrieval by using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).

Immunohistochemical analysis of paraffin-embedded rat liver tissue labeling ABAT/GABA-T with ab216465 at 1/500 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Granular cytoplasmic staining in rat liver (PMID: 25771305; PMID: 25738457) is observed. Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).

Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

Immunohistochemical analysis of paraffin-embedded mouse testis tissue labeling ABAT/GABA-T with ab216465 at 1/500 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Granular cytoplasmic staining in mouse testis (PMID: 25771305; PMID: 25738457) is observed. Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).

Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

Immunohistochemical analysis of paraffin-embedded human breast cancer tissue labeling ABAT/GABA-T with ab216465 at 1/500 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Granular cytoplasmic staining in human breast cancer (PMID: 25771305; PMID: 25738457) is observed. Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).

Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

Immunohistochemical analysis of paraffin-embedded human kidney tissue labeling ABAT/GABA-T with ab216465 at 1/500 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Granular cytoplasmic staining in human kidney (PMID: 25771305; PMID: 25738457) is observed. Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).

Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).