Anti-GFAP antibody [EPR1034Y] - Astrocyte marker - Rat IgG2a (Chimeric)

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GFAP antibody [EPR1034Y] - Astrocyte marker - Rat IgG2a (Chimeric) (AB279291)

IHC image of GFAP staining in a section of formalin-fixed paraffin-embedded normal human cerebral cortex 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 20mins. The section was then incubated with ab279291, 1μg/ml, for 15 mins at room temperature. A rabbit anti-rat IgG2a, ab102248, was added for 8 mins at room temperature and detected using an HRP conjugated goat anti-rabbit compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. The inset secondary-only control image is taken from an identical assay without primary antibody.
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.

• ICC

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Immunocytochemistry - Anti-GFAP antibody [EPR1034Y] - Astrocyte marker - Rat IgG2a (Chimeric) (AB279291)

Immunocytochemical analysis of 4% PFA-fixed, 0.1% Triton X-100 permeabilized mouse primary neural/glia tissue labeling GFAP with ab279291 at 1/50 (17 μg/ml) dilution followed by ab150161 Goat Anti-Rat IgG (Alexa Fluor® 488) pre-adsorbed at 1/1000 dilution (Green). Positive staining on mouse primary astrocytes is observed. The nuclear counterstain was DAPI (Blue).

Counterstain : ab207165 anti-GFAP rabbit mAb at 1/1000 dilution with secondary antibody ab150088 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) pre-adsorbed at 1/1000 dilution.

Negative controls : ab279291 at 1/50 dilution with secondary ab150080 at 1/500 dilution; and ab207165 at 1/1000 dilution with secondary ab150161 at 1/1000 dilution

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-GFAP antibody [EPR1034Y] - Astrocyte marker - Rat IgG2a (Chimeric) (AB279291)

Flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol-permeabilized rat primary neural/glia cells labelling GFAP with ab279291 at 1/1000 dilution (0.1μg)/ Right compared with a Rat monoclonal IgG isotype control/ Left.

Goat F(ab)2 Anti-Rat IgG Fc (Alexa Fluor® 647, ab150163) at 1/2000 dilution was used as the secondary antibody.

• IHC-Fr

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Immunohistochemistry (Frozen sections) - Anti-GFAP antibody [EPR1034Y] - Astrocyte marker - Rat IgG2a (Chimeric) (AB279291)

Immunohistochemical analysis of 4% PFA fixed 0.2% Triton X-100 permeabilized fresh frozen rat cerebrum tissue labeling GFAP with ab279291 at a 1/200 dilution, followed by Goat Anti-Rat IgG H&L (Alexa Fluor^® 488) (ab150157) secondary antibody at a 1/1000 dilution. The nuclear counterstain was DAPI (blue).

Positive staining on rat cerebrum.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rat IgG H&L (Alexa Fluor^® 488) ab150157 at a 1/1000 dilution.

• IHC-Fr

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Immunohistochemistry (Frozen sections) - Anti-GFAP antibody [EPR1034Y] - Astrocyte marker - Rat IgG2a (Chimeric) (AB279291)

Immunohistochemical analysis of 4% PFA fixed 0.2% Triton X-100 permeabilized fresh frozen mouse cerebrum tissue labeling GFAP with ab279291 at a 1/200 dilution, followed by Goat Anti-Rat IgG H&L (Alexa Fluor^® 488) (ab150157) secondary antibody at a 1/1000 dilution. The nuclear counterstain was DAPI (blue).

Positive staining on mouse cerebrum.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rat IgG H&L (Alexa Fluor^® 488) ab150157 at a 1/1000 dilution.

• WB

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Western blot - Anti-GFAP antibody [EPR1034Y] - Astrocyte marker - Rat IgG2a (Chimeric) (AB279291)

The molecular weight observed is consistent with the literature (PMID : 824020, 2294, 6340792).

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure times : Lane 1 : 3.25 seconds; Lane 2, 3 : 10 seconds.

All lanes:

Western blot - Anti-GFAP antibody [EPR1034Y] - Astrocyte marker - Rat IgG2a (Chimeric) (ab279291) at 1/1000 dilution

Lane 1:

Human brain tissue lysate at 20 µg

Lane 2:

Mouse brain tissue lysate at 20 µg

Lane 3:

Rat brain tissue lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rat IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rat-igg-h-l-hrp-ab205720'>ab205720</a>) at 1/5000 dilution

Predicted band size: 49 kDa

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