Recombinant Anti-GFAP antibody [EP672Y] (ab33922)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized Mouse primary brain cells cells labelling GFAP with ab33922 at 1/500 dilution (0.1ug)/ Right compared with a Rabbit monoclonal IgG (ab172730) / Left isotype control . A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.

ab33922 showing positive staining in Normal brain tissue.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

Immunofluorescence staining of GFAP using ab33922 in primary rat hippocampal mixed glia, (prepared from P2 rat hippocampal brain area, obtained from Transnetyx Tissue by BrainBits, LLC, cat.no. SDPHP4m), DIV4. The cells were fixed with 100% MeOH (5 min), permeabilized with 0.1% Triton-X-100 (in PBS) for 5 mins and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab33922 at 0.2 μg/ml and ab4674, Anti-GFAP antibody, at 1/1000 dilution. Cells were then incubated with ab150081, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (shown in green) and ab150176, Goat Anti-Chicken IgY H&L (Alexa Fluor® 594) preadsorbed at 1/1000 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).

Images were acquired with the Perkin Elmer Operetta HCA and a maximum intensity projection of confocal sections is shown. The antibody ab33922 gave comparable results using 4% formaldehyde fixation (10 min).

Immunocytochemistry analysis of Embryonic mouse primary neural cells labeling GFAP with purified ab33922 at 1:100 dilution (10 μg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. 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. Cells were counterstained with Tau Mouse mAb 1:100 (5 μg/ml). ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) was used as the secondary antibody for the counter stain at 1:1000 (2 μg/ml) dilution. DAPI (blue) was used as nuclear counterstain. 

Confocal image showing positive staining in glial cells.

ICC image of ab33922 stained Rat primary mixed astrocytes culture. The cells were 100% Paraformaldehyde fixed and then incubated in 10% Serum / 0.1M PBS with 10% Donkey serum for 4h. The secondary antibody was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution.

ab33922 showing positive staining in Astrocytoma tissue.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

ab33922 showing negative staining in Meningioma tissue.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.