重组Anti-GAD65 + GAD67抗体[EPR19366]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GAD65 + GAD67 antibody [EPR19366] (AB183999)

Immunohistochemical analysis of paraffin-embedded Mouse cerebellum tissue labeling GAD65 + GAD67 with ab183999 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Cytoplasm staining on mouse cerebellum is observed. Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is ab97051 at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IP

Supplier Data

Immunoprecipitation - Anti-GAD65 + GAD67 antibody [EPR19366] (AB183999)

ab183999 Immunoprecipitating GAD65 + GAD67 in Human cerebellum lysate. 10μg of cell lysate was incubated with primary antibody 1/40. For western blotting ab183999 (1/1000) was used to confirm successful immunoprecipation. VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/5000.

Lane 1 (Input) : Human cerebellum lysate 10μg
Lane 2 (+) : Human cerebellum lysate with ab183999, 1/40
Lane 3 (-) : Rabbit monoclonal IgG (ab172730) instead of ab183999 in Human cerebellum lysate

All lanes:

Immunoprecipitation - Anti-GAD65 + GAD67 antibody [EPR19366] (ab183999)

Predicted band size: 65 kDa

false

Exposure time: 2min

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GAD65 + GAD67 antibody [EPR19366] (AB183999)

Immunohistochemical analysis of paraffin-embedded Rat pancreas tissue labeling GAD65 + GAD67 with ab183999 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Cytoplasm staining on rat pancreas islet is observed. Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is ab97051 at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GAD65 + GAD67 antibody [EPR19366] (AB183999)

Immunohistochemical analysis of paraffin-embedded rat kidney tissue labeling GAD65 + GAD67 with ab183999 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Negative on rat kidney. Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is ab97051 at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GAD65 + GAD67 antibody [EPR19366] (AB183999)

Immunohistochemical analysis of paraffin-embedded Mouse lung tissue labeling GAD65 + GAD67 with ab183999 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Negative on mouse lung. Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is ab97051 at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GAD65 + GAD67 antibody [EPR19366] (AB183999)

Immunohistochemical analysis of formalin fixed paraffin embedded mouse cerebellum labelling GAD65 + GAD67 with ab183999 at a concentration of 0.1 μg/ml. The immunostaining was performed on a Ventana DISCOVERY ULTRA (Roche Tissue Diagnostics) instrument with a ChromoMap DAB Kit with anti-rabbit HQ and anti-HQ HRP detection. The heat-mediated antigen retrieval was conducted with DISCOVERY cell conditioning solution (CC1) at 100 °C, pH 8.5 for 32 mins. ab183999 Anti-GAD65 + GAD67 antibody [EPR19366] was incubated for 16 mins, at 37 °C. Sections were counterstained with Hematoxylin II. Image inset shows absence of staining in secondary antibody only control. Customers are encouraged to optimise antigen retrieval conditions, antibody concentration, incubation times and temperature for best results in their own IHC assay workflow (automated and manual).

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GAD65 + GAD67 antibody [EPR19366] (AB183999)

Immunohistochemical analysis of paraffin-embedded Rat cerebellum tissue labeling GAD65 + GAD67 with ab183999 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Cytoplasm staining on rat cerebellum is observed. Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is ab97051 at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-Fr

Supplier Data

Immunohistochemistry (Frozen sections) - Anti-GAD65 + GAD67 antibody [EPR19366] (AB183999)

Immunohistochemical analysis of 4% paraformaldehyde fixed, 0.2% Triton X-100 permealized frozen section of Mouse hippocampus tissue labeling GAD65 + GAD67 with ab183999 at 1/250 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor^® 488) secondary (ab150077) at 1/1000 dilution (green). The result showed mainly cytoplasmic staining on mouse hippocampus. The nuclear counterstain is DAPI (blue).

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is ab150077 at 1/1000 dilution.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-GAD65 + GAD67 antibody [EPR19366] (AB183999)

Immunocytochemistry/ Immunofluorescence analysis of mouse primary neuron cells labeling GAD65 + GAD67 with purified ab183999 at 1/100 (10 μg/mL). Cells were fixed in 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. Cells were counterstained with ab11267 Anti-MAP2 antibody [HM-2]; ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) was used as the secondary antibody at 1/1000 (2 μg/mL) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.
Confocal scanning Z step was set as 0.3 μm followed by image processing with maximum Z projection.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GAD65 + GAD67 antibody [EPR19366] (AB183999)

Immunohistochemical analysis of paraffin-embedded Mouse pancreas tissue labeling GAD65 + GAD67 with ab183999 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Cytoplasm staining on mouse pancreas islets is observed. Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is ab97051 at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• WB

Supplier Data

Western blot - Anti-GAD65 + GAD67 antibody [EPR19366] (AB183999)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure time : Lane 1-5 : 3 minutes; Lane 6 and 7 : 2 seconds.

All lanes:

Western blot - Anti-GAD65 + GAD67 antibody [EPR19366] (ab183999) at 1/1000 dilution

Lane 1:

Human cerebellum lysate at 20 µg

Lane 2:

Mouse skin lysate at 20 µg

Lane 3:

Mouse lung lysate at 20 µg

Lane 4:

Neuro-2a (Mouse neuroblastoma cell line) whole cell lysate at 20 µg

Lane 5:

C6 (Rat glial tumor cell line) whole cell lysate at 20 µg

Lane 6:

Mouse cerebellum lysate at 20 µg

Lane 7:

Rat cerebellum lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 65 kDa

Observed band size: 65 kDa,67 kDa

false

• WB

Supplier Data

Western blot - Anti-GAD65 + GAD67 antibody [EPR19366] (AB183999)

Blocking/Dilution buffer : 5% NFDM/TBST.

Lane 1:

Western blot - Anti-GAD65 + GAD67 antibody [EPR19366] (ab183999) at 1/5000 dilution

Lane 2:

Western blot - Anti-GAD65 + GAD67 antibody [EPR19366] (ab183999) at 1/1000 dilution

Lane 1:

Mouse brain lysate at 10 µg

Lane 2:

Rat brain lysate at 10 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 65 kDa

Observed band size: 65 kDa,67 kDa

false

Exposure time: 2s

• WB

Supplier Data

Western blot - Anti-GAD65 + GAD67 antibody [EPR19366] (AB183999)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure time : Lane 1 : 1 second; Lane 2 : 15 seconds.

Mouse GAD67 fragment recombinant protein contain aa468-592 with a His-Tag®. Mouse GAD65 fragment recombinant protein contain aa460-584 with a His-Tag®. These two fragment recombinant proteins were made in-house. The ~30 kDa band represents doublets of the recombinant fragments.

All lanes:

Western blot - Anti-GAD65 + GAD67 antibody [EPR19366] (ab183999) at 1/1000 dilution

Lane 1:

Mouse GAD67 fragment recombinant protein at 0.01 µg

Lane 2:

Mouse GAD65 fragment recombinant protein at 0.01 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 65 kDa

false