PhosphoTracer AKT 1/2/3 (pS473) + total AKT 1/2/3 ELISA Kit (ab119654)
- Product namePhosphoTracer AKT 1/2/3 (pS473) + total AKT 1/2/3 ELISA KitSee all AKT 1/2/3 (phospho Ser473) + total AKT 1/2/3 kits ...
- Tests1 x 96 well plate
- Sample typeCell culture extracts
- Assay typeSandwich
- Assay time1h 15m
- Species reactivityReacts with: Mouse, Hamster, Human
Predicted to work with: Rat
- Product overview
PhosphoTracer assays use a traditional immuno-sandwich format, but with a major difference both the analyte and the assay reagents are added to the PhosphoTracer assay microplate at the same time. After a short incubation period, unbound assay reagents and analytes are washed away, and immuno-complexes containing both antibodies are detected. The process can take as little as 60 minutes to complete.
PhosphoTracer kits also allow a higher degree of assay flexibility. In contrast to other ELISA formats, no antibodies are present on the assay microplate itself, so assays for several different targets can be performed in different wells on the same microplate. Simply mix the lysate with your target reagents of choice, using the microplate configuration of your choice.
A whole new way of performing cellular assays, PhosphoTracer takes the hard work out of running a standard ELISA, while still giving the high quality results expected from a sandwich immunoassay. Fully self-contained kits are supplied in convenient 96-well packs. Simple to use and highly sensitive PhosphoTracer kits are designed to get results, fast.
Abcam’s PhosphoTracer AKT 1/2/3 assay kits detect endogenous levels of AKT 1/2/3 (GenBank Accessions NP_001014431 [AKT1], NP_001617 [AKT2], and NP_859029 [AKT3]) in cellular lysates. Phospho-AKT 1/2/3 assays only detect AKT when phosphorylated at Ser473. Total AKT assays detect AKT regardless of phosphorylation state.
The substrate used with the HRP conjugated detection antibody is a combination of 10-Acetyl-3,7-dihydroxyphenoxazine (ADHP) (wavelength exc/em = 530-540nm / 590-600nm), a highly sensitive and stable substrate for HRP) and ADHP Dilution Buffer (a stabilized H2O2 solution). Learn more about the fluorogenic substrate, ADHP.
Sensitivity: Phospho-AKT 1/2/3: 10 pg/ml (Tested with rhAKT1), AKT 1/2/3: 100 pg/ml (Tested with rhAKT1).
Range: Phospho-AKT 1/2/3: 10-10,000 pg/ml, AKT 1/2/3: 100-100,000 pg/ml.
- Tested applicationsSandwich ELISA more details
- Storage instructionsStore at +4°C. Please refer to protocols.
Components 1 x 96 tests 96-well PhosphoTracer assay plate (stripwell) 1 unit Adherent plate seal 2 units ADHP (100X) 1 x 120µl ADHP Dilution Buffer 1 x 15ml Assay Control Lysate (lyophilized) 1 x 0.25ml Enhancer Solution 1 x 1ml Lysis Buffer (5X) 1 x 15ml Mouse monoclonal AKT 1/2/3 (24 assay points) 1 x 0.75ml Mouse monoclonal Phospho-AKT (Ser473) (72 assay points) 1 x 3ml Rabbit monoclonal Phospho-AKT 1/2/3 (HRP) (72 assay points) 3 x 0.75ml Rabbit polyclonal AKT 1/2/3 (HRP) (24 assay points) 1 x 0.75ml Stop Solution 1 x 2ml Wash Buffer (10X) 1 x 15ml
- Research Areas
Contains 1 AGC-kinase C-terminal domain.
Contains 1 PH domain.
Contains 1 protein kinase domain.
modificationsPhosphorylation on Thr-309 and Ser-474 is required for full activity.
Ubiquitinated; undergoes both 'Lys-48'- and 'Lys-63'-linked polyubiquitination. TRAF6-induced 'Lys-63'-linked AKT2 ubiquitination. When fully phosphorylated and translocated into the nucleus, undergoes 'Lys-48'-polyubiquitination catalyzed by TTC3, leading to its degradation by the proteasome.
- AKT2_HUMANAKT3HIHGHHPKBPKB alphaPKB betaPKB gammaPKB-ALPHAPKB-GAMMAPKBBPKBBETAPKBGPRKBAPRKBBPRKBGProtein kinase Akt-2Protein kinase Akt-3Protein kinase BProtein kinase B alphaProtein kinase B betaProtein kinase B gammaProto oncogene c AktRACRAC-ALPHARAC-alpha serine/threonine-protein kinaseRAC-BETARAC-beta serine/threonine-protein kinaseRAC-gammaRAC-gamma serine/threonine-protein kinaseRAC-PK-alphaRAC-PK-betaRAC-PK-gammaSTK-2V-akt murine thymoma viral oncogene homolog 1V-akt murine thymoma viral oncogene homolog 2V-akt murine thymoma viral oncogene homolog 3 (protein kinase B, gamma)
- Entrez Gene: 208 Human
- Entrez Gene: 10000 Human
- Entrez Gene: 207 Human
- Entrez Gene: 11652 Mouse
- Entrez Gene: 11651 Mouse
- Entrez Gene: 23797 Mouse
- Entrez Gene: 25233 Rat
- Entrez Gene: 24185 Rat
- Entrez Gene: 29414 Rat
- Omim: 164731 Human
- Omim: 164730 Human
- Omim: 611223 Human
- SwissProt: P31751 Human
- SwissProt: P31749 Human
- SwissProt: Q9Y243 Human
- SwissProt: Q60823 Mouse
- SwissProt: P31750 Mouse
- SwissProt: Q9WUA6 Mouse
- SwissProt: P47197 Rat
- SwissProt: P47196 Rat
- SwissProt: Q63484 Rat
- Unigene: 631535 Human
Our Abpromise guarantee covers the use of ab119654 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|Sandwich ELISA||sELISA. Phospho-AKT 1/2/3: Tested in HEK293, HeLa, MCF7, Jurkat, CHO; AKT 1/2/3: Tested in HEK293, HeLa, MCF7, A431, C2C12, Raw264.7, A549, PC3, NIH3T3, Jurkat.|
PhosphoTracer AKT 1/2/3 (pS473) + total AKT 1/2/3 ELISA Kit images
Using Western blot, a significant increase in AKT 1/2/3 phosphorylation at Ser473 is detected in MCF7 cells treated with insulin for 15 minutes (+), compared with untreated cells (-), whereas no change in total AKT 1/2/3 levels was observed.
Using the AKT 1/2/3 assay kits, a significant increase in AKT 1/2/3 phosphorylation at Ser473 is detected in MCF7 cells treated with insulin for 15 minutes (+), compared with untreated cells (-), whereas no change in total AKT 1/2/3 levels was observed.
Using the PhosphoTracer phospho-AKT assay, the length of assay incubation time that was required to detect recombinant phospho-AKT was examined. After 15 mins, phospho-AKT concentrations of approximately 100 pg/mL were readily detected, while after 1 hour incubation, the limit of detection was less than 10 pg/mL. Longer incubation times had little effect on overall limit of detection that was observed.
Jurkat cells were harvested by centrifugation, resuspended in HBSS, and seeded at 10K/well (20 µL/well) in a rinsed PhosphoTracer assay plate. The cells were treated with various concentrations of wortmannin (10 µL/well) for 1 h at 37°C, then treated with 5% FCS (10 µL/well) for 15 min. The cells were lysed with 10 µL/well Lysis Mix Concentrate, with shaking for 10 min. Antibody Mix specific for phospho AKT (50 µL/well) was added to the lysate, and the plates were incubated for 1 hr at room temp, with shaking. Plates were washed and Substrate Mix was added. The plates were covered in foil and incubated for 10 min with shaking. Signal in the wells was determined using a plate reader.
References for PhosphoTracer AKT 1/2/3 (pS473) + total AKT 1/2/3 ELISA Kit (ab119654)
ab119654 has not yet been referenced specifically in any publications.