Recombinant Anti-GABA A Receptor alpha 6 antibody [EPR25322-129] (BSA and Azide free) (ab300070)

Fluorescence multiplex immunohistochemical analysis of human cerebellum (formalin-fixed paraffin-embedded section). Merged staining of ab300069, GABA A Receptor alpha 6 at 1:500 (1.244 μg/ml) [Panel B], ab239790, DGKZ/DGK-zeta at 1:9000 (0.109 μg/ml) [Panel C], and ab108404, Calbindin at 1:1000 (1.394 μg/ml) [Panel D] on human cerebellum. Followed by Opal Polymer HRP Ms + Rb secondary. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument with an Opal™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope. The section was incubated in three rounds of staining: in the order of ab300069, ab239790, and ab108404 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins was used. DAPI (dark blue) was used as a nuclear counter stain.

This data was developed using ab300069, the same antibody clone in a different buffer formulation.

All lanes : Anti-GABA A Receptor alpha 6 antibody [EPR25322-129] (ab300069) at 1/1000 dilution

Lane 1 : Human hypothalamus tissue lysate
Lane 2 : Human heart tissue lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : VeriBlot for IP Detection Reagent (HRP) (ab131366) at 1/1000 dilution

Predicted band size: 51 kDa
Observed band size: 57 kDa why is the actual band size different from the predicted?


Exposure time: 180 seconds

This data was developed using ab300069, the same antibody clone in a different buffer formulation. 

5% NFDM/TBST was used as blocking and diluting buffer. 

Low expression:Heart (PMID:29467616).

Anti-GABA A Receptor alpha 6 antibody [EPR25322-129] (ab300069) at 1/1000 dilution + Mouse brain tissue lysate at 20 µg

Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

Predicted band size: 51 kDa
Observed band size: 57 kDa why is the actual band size different from the predicted?


Exposure time: 59 seconds

This data was developed using ab300069, the same antibody clone in a different buffer formulation. 

5% NFDM/TBST was used as blocking and diluting buffer. 

All lanes : Anti-GABA A Receptor alpha 6 antibody [EPR25322-129] (ab300069) at 1/1000 dilution

Lane 1 : Rat brain tissue lysate
Lane 2 : Rat cerebellum tissue lysate
Lane 3 : Rat heart tissue lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG (Merck DC03L) at 1/2000 dilution

Predicted band size: 51 kDa


Exposure time: 180 seconds

This data was developed using ab300069, the same antibody clone in a different buffer formulation. 

5% NFDM/TBST was used as blocking and diluting buffer. 

Low expression:Heart (PMID:29467616)
This blot was developed using a high sensitivity ECL substrate.

This data was developed using ab300069, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded Mouse cerebellum tissue labelling GABA A Receptor alpha 6 with ab300069 at 1/500 dilution (1.244 µg/ml) followed by a ready to use LeicaDS9800 (BOND™ Polymer Refine Detection). Positive staining on mouse cerebellum. The section was incubated with ab300069 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (BOND™ Polymer Refine Detection).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins. 

Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (BOND™ Polymer Refine Detection) was used.

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

This data was developed using ab300069, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded Rat cerebellum tissue labelling GABA A Receptor alpha 6 with ab300069 at 1/500 dilution (1.244 µg/ml) followed by a ready to use LeicaDS9800 (BOND™ Polymer Refine Detection). Positive staining on rat cerebellum. The section was incubated with ab300069 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (BOND™ Polymer Refine Detection).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins. 

This data was developed using ab300069, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded Human cerebellum tissue labelling GABA A Receptor alpha 6 with ab300069 at 1/500 dilution (1.244 µg/ml) followed by a ready to use LeicaDS9800 (BOND™ Polymer Refine Detection) was used. Positive staining on human cerebellum. The section was incubated with ab300069 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument.  Counterstained with Hematoxylin.

Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (BOND™ Polymer Refine Detection).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins. 

This data was developed using ab300069, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Mouse cerebellum (fresh) tissue labeling GABA A Receptor alpha 6 with ab300069 at 1/500 (1.244 µg/ml) dilution followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed at 1/1000 2 µg/mL dilution (Green). Positive staining on mouse cerebellum is observed. The nuclear counterstain was DAPI (Blue).

Secondary antibody control: Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbedat 1/1000 2 µ/mL dilution.

This data was developed using ab300069, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Rat cerebellum (fresh) tissue labeling GABA A Receptor alpha 6 with ab300069 at 1/500 (1.244 µg/ml) dilution followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed at 1/1000 (2 µg/mL) dilution (Green). Positive staining on rat cerebellum is observed. The nuclear counterstain was DAPI (Blue).

Secondary antibody control: Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbedat 1/1000 (2 µ/mL) dilution.