GFAP Antibody | anti-GFAP antibody

GFAP, Monoclonal Antibody, Mouse

Cat. Num. AAA76413

• Gene Names: GFAP; ALXDRD
• Reactivity: Human, Rat, Mouse, Primate, Cat, Cow, Pig, Horse, Chicken
• Applications: Western Blot, Immunohistochemistry, Immunofluorescence
• Purity: Purified
• Synonyms: GFAP; N/A; GFAP, Monoclonal Antibody, Mouse; anti-GFAP antibody

• Reactivity: Human, Rat, Mouse, Primate, Cat, Cow, Pig, Horse, Chicken
• Isotype: IgG1, with a k light chain
• Purity/Purification: Purified
• Form/Format: Liquid with 10mM Sodium Azide
• Sequence Length: 432

• Applicable Applications for anti-GFAP antibody: WB (Western Blot), IHC (Immunohistochemistry), IF (Immunofluorescence)
• Immunostaining Cell Cultures: 1. Draw of culture medium with aspirator and add 1 ml of 3.7 % formalin in PBS solution to the dish. (make up from 10mls Fisher 37% formalin plus 90mls PBS, the Fisher formalin contains 37% formaldehyde plus about 1% methanol which may be relevant sometimes). Let sit at room temp for 1 minute. (can add 0.1% Tween 20 to PBS used here and all subsequent steps to reduce background; probably best not to do this first time round though as it may extract your antigen or help wash your cells off the dish).; 2. Take off the formalin/PBS and add 1ml of cold methanol (-20°C, kept in well sealed bottle in fridge). Let sit for no more than 1 minute.; 3. Take off methanol and add 1ml of PBS, not letting the specimen dry out. To block nonspecific antibody binding can add ~10ml (=1%) of goat serum (Sigma), and can incubate for 30 minutes. Can then add antibody reagents. Typically 100ml of hybridoma tissue culture supernatent or 1ml of mouse ascites fluid or crude serum. Incubate for 1 hour at room temp. (or can go at 37°C for 30 minutes to 1 hour, or can do 4°C overnight, exact time not too critical). Can do very gentle shaking for well adherent cell lines (3T3, Hek293 etc.).; 4. Remove primary antibody and replace with 1 ml of PBS. Let sit for 5-10 minutes, replace PBS and repeat twice, to give three washes in PBS.; 5. Add 0.5 mls of secondary antibody. These are fluorescently labeled Goat anti mouse or rabbit antibodies and are conjugated to ALEXA dyes and are from Molecular probes (Eugene Oregon, the ALEXA dyes are sulphonated rhodamine compounds and are much more stable to UV than FITC, TRITC, Texas red etc.). Typically make 1:2,000 dilutions of these secondaries in PBS plus 1% goat serum, BSA or non fat milk carrier. Incubate for 1 hour at room temp. (or can go at 37°C for 30 minutes to 1 hour, or can do 4°C overnight). Can do gentle shaking for well adherent cell lines (3T3, HEK293 etc.).; 6. Remove secondary antibody and replace with 1 ml of PBS. Let sit for 5-10 minutes, replace PBS and repeat twice, to give three washes in PBS. ; 7. Drop on one drop of Fisher mounting medium onto dish and apply 22mm square coverslip. View in the microscope
• Immunostaining Tissue: Solutions: PBS - sodium phosphate-buffered (100 mM; pH 7.2) isotonic (0.9% NaCl, w/v) saline Antibody dilution buffer (PBS with 0.1% non-ionic detergent, such as Triton X-100 or Tween-20) fluorescein anti-fading reagent -- Make up a 2 mg/ml phenylene diamine solution in PBS (phenylene diamine requires extensive vortexing to put it into solution). Once the phenylene diamine is completely dissolved, add an equal volume of glycerol and mix. This reagent will last about a week at -20°C. Discard this reagent when it starts to turn dark brown. Other Reagents Fluorescein-labeled goat anti-mouse IgG; 1. Prepare your tissue sections or cultured cells as you normally would. Wash your sections or cells for 1 min with PBS at room temperature.; 2. Incubate your sections or cells with your chicken primary antibodies (diluted in "antibody dilution buffer") for at least 1 hour at room temperature. The concentration of your antibody may be anywhere from 1:50-1:150 depending on the titre of the antibody and the concentration of your antigen.; 3. Wash your sections or cells over a 10 minute period at room temperature (with two changes of PBS).; 4. Incubate your sections or cells with fluorescein-labeled goat anti-mouse IgG (1:500 dilution in "antibody dilution buffer” for 1 hour at room temperature. Be sure to keep these slides or culture dishes in subdued light (e.g., in a drawer) to avoid bleaching of the fluorescein dye.; 5. Repeat step #4; 6. Add a drop of "fluorescence anti-fading reagent" (i-BRITE Plus) to your sections or cells. Place a coverslip over the section. If you want to reduce messiness, you may also seal the coverslip by painting the edges with nail polish.; 7. Store the slides or culture dishes in the refrigerator (in the dark).
• Immunogen Sequence: Recombinant GFAP expressed in bacteria and highly purified. Subsequent boosts were performed with GFAP purified from a Triton X-100 extract of myelin associated material from bovine spinal cord.
• Preparation and Storage: Antibody can also be aliquotted and stored frozen at -20° C to -70° C in a manual defrost freezer for six months without detectable loss of activity.; The antibody can be stored at 2° - 8° C for 1 month without detectable loss of activity. Avoid repeated freeze-thaw cycles.

Application Data

(Image: Strip blot of rat spinal cord protein extract stained with GFAP. A prominent band at about 55 kDa corresponds to the major isoform of GFAP.)

Application Data

(Images: Mixed neuron-glial cultures stained with Mouse Monoclonal GFAP, and Chicken Polyclonal Neurofilament-NF-L (green). The GFAP antibody stains the network of astrocytes in these cultures, while the NF-L antibody stains neurons and their processes. The blue channel shows the localization of DNA. This antibody also works on formalin fixed paraffin embedded brain tissues. Protocol on Datasheet.)

Product Categories/Family for anti-GFAP antibody

Antibodies; Neuronal-Glial Markers; Glial-Astrocyte Markers; GFAP

NCBI and Uniprot Product Information

• NCBI GI #: 121135
• NCBI GeneID: 2670
• NCBI Official Full Name: Glial fibrillary acidic protein
• NCBI Official Synonym Full Names: glial fibrillary acidic protein
• NCBI Official Symbol: GFAP
• NCBI Official Synonym Symbols: ALXDRD
• NCBI Protein Information: glial fibrillary acidic protein
• UniProt Protein Name: Glial fibrillary acidic protein
• UniProt Gene Name: GFAP
• UniProt Synonym Gene Names: GFAP
• UniProt Entry Name: GFAP_HUMAN