IHC-P image using GTX24802- Dectection of AKT by IHC-P in human breast carcinoma tissue
|
Double immunofluorescent staining of phospho-AKT Ser 473 (GTX24802; green) and beta III tubulin (red) in [A] non-stimulated and [B] NGF stimulated adult DRG neurons. Following two days in culture, neurons were stimulated [B] with NGF (100ng/ml for 10 min at 37°C). NGF treatment induced an increased phosphorylation of p-AKT Ser 473 (GTX24802; [B] increased green staining) in NGF sensitive DRG neurons. beta III tubulin is neuronal marker used to visualise DRG neurons. Image courtesy of Dr S Pezet, CARD institute, KCL, London, UK
|
Immunofluorescence detection of AKT phospho S473 antibody (GTX24802) at a dilution of 1/200 (24h incubation at RT; Alexa fluor 488 secondary antibody 1/1000). Perinuclear staining observed in dorsal and ventral horn neurons of rat spinal cord (40x objective). Immunofluorescence staining performed on slide with 30µm saggital spinal cord sections.
|
Dectection of AKT by WB in human breast carcinoma tissue
|
Peptide Competition: Extracts of NIH-3T3 cells stimulated with PDGF were resolved by SDS-PAGE on a 10% polyacrylamide gel and transferred to nitrocellulose. Membranes were blocked with a 4% BSA-TBST buffer overnight at 4°C, then were incubated with 0.50 µg/mL ab4802 for 2h at RT in 3% BSA-TBST, following, prior incubation with: the phosphopeptide immunogen (1), a generic phosphoserine-containing peptide (2), the non-phosphorylated peptide corresponding to the immunogen (3), or, no peptide (4). After washing, membranes were incubated with secondary goat anti-rabbit IgG alkaline phosphatase and bands were detected using the Tropix WesternStar detection method. The data show that only the phosphopeptide corresponding to GT4802 completely blocks the antibody signal, demonstrating the specifici
|
|
|
|
Product Name
AKT (phospho S473) antibody
Catalog Number
GTX24802
Product Description:
Rabbit polyclonal to AKT ( phospho S473 Rabbit polyclonal to AKT ( phospho S473 )
Application Note:
IHC-P: Use at an assay dependent dilution. WB: Use at a dilution of 1/500 - 1/1000. Predicted molecular weight: 65 kDa. Can be blocked with GTX25308 AKT peptides (phospho S473) (phospho and non-phospho pair) . Not tested in other applications. Optimal dilutions/concentrations should be determined by the end user.
Tested Applications:
IHC (Formalin-fixed paraffin-embedded sections), Western blot. The usefulness of this product in other applications has not been determined.
Background:
AKT, also known as Protein Kinase B (PKB) or RAC alpha, is a 65 kDa serine/threonine kinase that plays an important role in many biological responses including metabolism, cell survival and growth by phosphorylating many proteins including GSK 3 beta, caspase 9, BAD, and the Forkhead Transcription Factor. AKT is activated by PI3 K, which in turn can interact with proteins such as Focal Adhesion Kinase (when Focal Adhesion Kinase is phosphorylated at Tyr397), thereby linking activation to the cytoskeleton. AKT is phosphorylated on Thr308 by PDK 1 and on Ser473 by PDK 2. Phosphorylation at Ser473 is required for full activation of AKT.
Clonality:
Polyclonal
Specificity:
Reacts with Human and mouse AKT 1, 2 and 3
Immunogen:
Synthetic peptide derived from the region of human AKT that contains serine 473. The sequence is conserved among multiple species including mouse and rat across the three isoforms, AKT1, AKT2, and AKT3.
Host:
Rabbit
Isotype:
IgG
Cross Reactivity:
Human, Mouse, and Rat - Not yet tested in other species - Expected to cross-react with a wide range of other species due to sequence homology
Positive Control:
NIH3 T3 cells stimulated with PDGF and 3T3 L1 adipocytes stimulated with OSM and IFN gamma
Target Protein:
AKT
Cellular Localization:
Cytoplasmic and Nuclear after activation by integrin linked protein kinase 1 (ILK1)
Form:
Liquid
Concentration:
Batch dependent
Purity:
Affinity purified with antigen
Purification Note:
The antibody was affinity-purified from rabbit antiserum by affinity-chromatography using epitope-specific phosphopeptide. The antibody against non-phosphopeptide was removed by chromatogramphy using non-phosphopeptide corresponding to the phosphorylation site.
Storage Buffer:
Phosphate-buffered saline, pH 7.3, 50% Glycerol and 150mM NaCl containing 0.02% sodium azide
Storage Instruction:
Upon Receipt - Keep as concentrated solution. Aliquot and store at -20ºC or below. Avoid freeze-thaw cycles.
Specific References:
1. Bremer K [Chronic lymphatic leukemia: disorders of lymphocyte kinetics. Pathophysiological aspects and therapeutic consideration] Dtsch Med Wochenschr 100:1250-5 (1975).
1975
1132340
2. Schwartzbauer G & Robbins J The tumor suppressor gene PTEN can regulate cardiac hypertrophy and survival. J Biol Chem 276:35786-93 (2001).
2001
11448956
3. Hsu J et al. The AKT kinase is activated in multiple myeloma tumor cells. Blood 98:2853-5 (2001).
2001
11675360
4. Morel JC et al. Signal transduction pathways involved in rheumatoid arthritis synovial fibroblast interleukin-18-induced vascular cell adhesion molecule-1 expression. J Biol Chem 277:34679-91 (2002).
2002
12105209
5. Beffert U et al. Reelin-mediated signaling locally regulates protein kinase B/Akt and glycogen synthase kinase 3beta. J Biol Chem 277:49958-64 (2002).
2002
12376533
|
| Please tell us about your experience or submit a specific inquiry about this product. |
|
|
|
|
|
