Western blot analysis of GFAP using anti-GFAP antibody (PB9082).
Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 30 ug of sample under reducing conditions.
Lane 1: human U251 whole cell lysates,
Lane 2: human Hela whole cell lysates,
Lane 3: rat brain tissue lysates,
Lane 4: rat C5 whole cell lysates,
Lane 5: mouse brain tissue lysates.
After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. Blocked the membrane with 5% non-fat milk/TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-GFAP antigen affinity purified polyclonal antibody (Catalog # PB9082) at 0.5 μg/mL overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:5000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1002) with Tanon 5200 system. A specific band was detected for GFAP at approximately 50 kDa. The expected band size for GFAP is at 50 kDa.

IF analysis of GFAP using anti-GFAP antibody (PB9082).
GFAP was detected in paraffin-embedded section of mouse brain tissues. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution ) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/mL rabbit anti-GFAP Antibody (PB9082) overnight at 4°C. Cy3 Conjugated Goat Anti-Rabbit IgG (BA1032) was used as secondary antibody at 1:100 dilution and incubated for 30 minutes at 37°C. The section was counterstained with DAPI. Visualize using a fluorescence microscope and filter sets appropriate for the label used.