AKT1 + AKT2 + AKT3 (pS473) ELISA Kit (ab176635)
Key features and details
- One-wash 90 minute protocol
- Sensitivity: 30 pg/ml
- Range: 1 ng/ml - 100 ng/ml
- Sample type: Cell Lysate, Tissue Homogenate
- Detection method: Colorimetric
- Assay type: Semi-quantitative
- Reacts with: Mouse, Human
Overview
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Product name
AKT1 + AKT2 + AKT3 (pS473) ELISA Kit
See all AKT1 + AKT2 + AKT3 kits -
Detection method
Colorimetric -
Precision
Intra-assay Sample n Mean SD CV% HEK lysate 6 = 5.4% Inter-assay Sample n Mean SD CV% HEK lysate 3 = 2.7% -
Sample type
Cell Lysate, Tissue Homogenate -
Assay type
Semi-quantitative -
Sensitivity
30 pg/ml -
Range
1 ng/ml - 100 ng/ml -
Recovery
4.2 %
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Assay time
1h 30m -
Assay duration
One step assay -
Species reactivity
Reacts with: Mouse, Human
Predicted to work with: Rat
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Product overview
Abcam’s AKT 1/2/3 (pS473) in vitro SimpleStep ELISA™ (Enzyme-Linked Immunosorbent Assay) kit is designed for the semi-quantitative measurement of AKT 1/2/3 (pS473) protein in Human and mouse cells.
The SimpleStep ELISA™ employs an affinity tag labeled capture antibody and a reporter conjugated detector antibody which immunocapture the sample analyte in solution. This entire complex (capture antibody/analyte/detector antibody) is in turn immobilized via immunoaffinity of an anti-tag antibody coating the well. To perform the assay, samples or standards are added to the wells, followed by the antibody mix. After incubation, the wells are washed to remove unbound material. TMB substrate is added and during incubation is catalyzed by HRP, generating blue coloration. This reaction is then stopped by addition of Stop Solution completing any color change from blue to yellow. Signal is generated proportionally to the amount of bound analyte and the intensity is measured at 450 nm. Optionally, instead of the endpoint reading, development of TMB can be recorded kinetically at 600 nm.
As of October 2019, this kit was reformulated with new antibodies to maintain continued long term supply.
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Notes
Abcam has not and does not intend to apply for the REACH Authorisation of customers’ uses of products that contain European Authorisation list (Annex XIV) substances.
It is the responsibility of our customers to check the necessity of application of REACH Authorisation, and any other relevant authorisations, for their intended uses. -
Platform
Microplate
Properties
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Storage instructions
Store at +4°C. Please refer to protocols. -
Components 1 x 96 tests 10X Wash Buffer PT 1 x 15ml 50X Cell Extraction Enhancer Solution 1 x 1ml 5X Cell Extraction Buffer PTR 1 x 12ml AKT 1/2/3 (pS473) Capture Antibody 1 x 3ml AKT 1/2/3 (pS473) Detector Antibody 1 x 3ml Lyophilized AKT Control Lysate 1 vial Plate Seal 1 unit SimpleStep Pre-Coated 96-Well Microplate (ab206978) 1 unit Stop Solution 1 x 12ml TMB Substrate 1 x 12ml -
Research areas
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Function
IGF-1 leads to the activation of AKT3, which may play a role in regulating cell survival. Capable of phosphorylating several known proteins. Truncated isoform 2/PKB gamma 1 without the second serine phosphorylation site could still be stimulated but to a lesser extent. -
Tissue specificity
In adult tissues, it is highly expressed in brain, lung and kidney, but weakly in heart, testis and liver. In fetal tissues, it is highly expressed in heart, liver and brain and not at all in kidney. -
Sequence similarities
Belongs to the protein kinase superfamily. AGC Ser/Thr protein kinase family. RAC subfamily.
Contains 1 AGC-kinase C-terminal domain.
Contains 1 PH domain.
Contains 1 protein kinase domain. -
Domain
Binding of the PH domain to the phosphatidylinositol 3-kinase alpha (PI(3)K) results in its targeting to the plasma membrane. -
Post-translational
modificationsPhosphorylation on Thr-305 and Ser-472 is required for full activity (By similarity). Phosphorylated upon DNA damage, probably by ATM or ATR.
Ubiquitinated. When fully phosphorylated and translocated into the nucleus, undergoes 'Lys-48'-polyubiquitination catalyzed by TTC3, leading to its degradation by the proteasome. -
Cellular localization
Cytoplasm. Membrane. Membrane-associated after cell stimulation leading to its translocation. - Information by UniProt
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Alternative names
- AKT
- AKT1
- AKT1 kinase
see all -
Database links
- Entrez Gene: 208 Human
- Entrez Gene: 207 Human
- Entrez Gene: 10000 Human
- Entrez Gene: 11651 Mouse
- Entrez Gene: 11652 Mouse
- Entrez Gene: 23797 Mouse
- Entrez Gene: 24185 Rat
- Entrez Gene: 25233 Rat
see all
Images
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Other - AKT 1/2/3 (pS473) ELISA Kit (ab176635)
SimpleStep ELISA technology allows the formation of the antibody-antigen complex in one single step, reducing assay time to 90 minutes. Add samples or standards and antibody mix to wells all at once, incubate, wash, and add your final substrate. See protocol for a detailed step-by-step guide.
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Typical Recombinant Protein Standard Curve -
Typical cell lysate dilution seriesExample of a typical AKT (pS473) cell lysate dilution series. Background-subtracted data values (mean +/- SD) are graphed.
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Linearity of dilutionLinearity of dilution in representative sample matrices. Cellular lysates were prepared at 3 concentrations in common media containing 1X Cell Extraction Buffer PTR. Data from duplicate measurements of AKT1 (pS473) are normalized and plotted.
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AKT (pS473) phosphorylation in response to insulin treatment.Induction of AKT (pS473) phosphorylation in MCF-7 cells in response to insulin treatment. MCF-7 cells were cultured in 96-well tissue culture plates, serum-starved and treated (5 min) with a dose-range of insulin before cell lysis. Data from quadruplicate measurements of AKT (pS473) are plotted and compared against total AKT1 protein levels. Comparative AKT (pS473) and AKT1 (Total) data also shown by Western Blot.
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Comparison of total AKT1 expression in different cell linesCell line analysis for total AKT1 from 20 µg/mL preparations of cell extracts. Data from triplicate measurements (mean +/- SD) are plotted and compared to 1X Cell Extraction Buffer PTR (zero).
