Human FOXO3 (FOXO3A) knockout HEK-293 cell lysate (ab261649)
Overview
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Product name
Human FOXO3 (FOXO3A) knockout HEK-293 cell lysate
See all FOXO3A kits -
Product overview
Knockout cell lysate achieved by CRISPR/Cas9.
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Parental Cell Line
HEK-293 -
Organism
Human -
Mutation description
Knockout achieved by CRISPR/Cas9; X = 15 bp deletion, 3 bp deletion -
Passage number
<20 -
Knockout validation
Next Generation Sequencing (NGS), Western Blot (WB) -
Reconstitution notes
To use as WB control, resuspend the lyophilizate in 50 µL of LDS* Sample Buffer to have a final concentration of 2 mg/ml. For reducing conditions, we recommend a final concentration of 0.1 M DTT.*Usage of SDS sample buffer is not recommended with these lyophilized lysates.
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Notes
Lysate preparation: Our lysates are made using RIPA buffer to which we add a protease inhibitor cocktail and phosphatase inhibitor cocktail (ratio: 300:100:10). This means that the protein of interest is denatured. If you require a native form of the protein please use the live cell version - found here. Please refer to our lysis protocol for further details on how our lysates are prepared.
User storage instructions: Lyophilizate may be stored at 4°C. After reconstitution, store at -20°C for short-term storage or -80°C for long-term storage.
Access thousands of knockout cell lysates, generated from commonly used cancer cell lines.
See here for more information on knockout cell lysates.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.This product is subject to limited use licenses from The Broad Institute and ERS Genomics Limited, and is developed with patented technology. For full details of the limited use licenses and relevant patents please refer to our limited use license and patent pages.
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Tested applications
Suitable for: WBmore details
Properties
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Storage instructions
Store at -80°C. Please refer to protocols. -
Components 1 kit ab280399 - Human FOXO3 knockout HEK293 cell lysate 1 x 100µg ab259780 - Human wild-type HEK-293 cell lysate 1 x 100µg -
Research areas
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Cell type
epithelial -
Gender
Female
Target
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Function
Transcriptional activator which triggers apoptosis in the absence of survival factors, including neuronal cell death upon oxidative stress. Recognizes and binds to the DNA sequence 5'-[AG]TAAA[TC]A-3'. -
Tissue specificity
Ubiquitous. -
Involvement in disease
Note=A chromosomal aberration involving FOXO3 is found in secondary acute leukemias. Translocation t(6;11)(q21;q23) with MLL/HRX. -
Sequence similarities
Contains 1 fork-head DNA-binding domain. -
Post-translational
modificationsIn the presence of survival factors such as IGF-1, phosphorylated on Thr-32 and Ser-253 by AKT1/PKB. This phosphorylated form then interacts with 14-3-3 proteins and is retained in the cytoplasm. Survival factor withdrawal induces dephosphorylation and promotes translocation to the nucleus where the dephosphorylated protein induces transcription of target genes and triggers apoptosis. Although AKT1/PKB doesn't appear to phosphorylate Ser-315 directly, it may activate other kinases that trigger phosphorylation at this residue. Phosphorylated by STK4 on Ser-209 upon oxidative stress, which leads to dissociation from YWHAB/14-3-3-beta and nuclear translocation. Phosphorylated by PIM1. -
Cellular localization
Cytoplasm > cytosol. Nucleus. Translocates to the nucleus upon oxidative stress and in the absence of survival factors. - Information by UniProt
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Alternative names
- AF6q21
- AF6q21 protein
- DKFZp781A0677
see all
Associated products
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KO cell lines
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab261649 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Application | Abreviews | Notes |
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WB |
Use at an assay dependent concentration.
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Notes |
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WB
Use at an assay dependent concentration. |
Images
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Lane 1: Wild-type HEK-293T cell lysate 20 ug
Lane 2: FOXO3 knockout HEK-293T cell lysate (ab256922) 20 ug
Lane 3: Wild-type HEK-293 cell lysate 20 ug
Lane 4: FOXO3 knockout HEK-293 cell lysate (ab261649) 20 ug
Lanes 1 - 4:Merged signal (red and green). Green - ab109629 observed at 82 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.
ab109629 was shown to react with FOXO3A in wild-type HEK-293 cells in Western blot with loss of signal observed in FOXO3 knockout cell line ab260857 (FOXO3 knockout cell lysate ab261649). Wild-type HEK-293 and FOXO3 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with ab109629 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4 °C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging. -
Lane 1: Wild-type HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate 20 ug
Lane 2: FOXO3 knockout HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate 20 ug
Lane 3: HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate 20 ug
Lane 4: Jurkat (Human T cell leukemia cell line from peripheral blood) whole cell lysate 20 ug
Lanes 1 - 4: Merged signal (red and green). Green - ab23683 observed at 71 kDa. Red - loading control, ab8245, observed at 37 kDa.
ab23683 was shown to recognize FOXO3A in wild-type HEK-293 cells as signal was lost at the expected MW in FOXO3 knockout cell line ab260857 (knockout cell lysate ab261649). Additional cross-reactive bands were observed in the wild-type and knockout samples. Wild-type and FOXO3 knockout samples were subjected to SDS-PAGE. Ab23683 and ab8245 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 1 µg/ml and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging. -
Lane 1: Wild-type HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate 20 ug
Lane 2: FOXO3 knockout HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate 20 ug
Lane 3: HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate 20 ug
Lane 4: Jurkat (Human T cell leukemia cell line from peripheral blood) whole cell lysate 20 ug
Lanes 1 - 4: Merged signal (red and green). Green - ab109629 observed at 71 kDa. Red - loading control, ab8245, observed at 37 kDa.
ab109629 was shown to recognize FOXO3A in wild-type HEK-293 cells as signal was lost at the expected MW in FOXO3 knockout cell line ab260857 (knockout cell lysate ab261649). Additional cross-reactive bands were observed in the wild-type and knockout cell lysate. Wild-type and FOXO3 knockout samples were subjected to SDS-PAGE. The membrane was blocked with 3% milk. Ab109629 and ab8245 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.
Protocols
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
Datasheets and documents
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SDS download
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Datasheet download
References (0)
ab261649 has not yet been referenced specifically in any publications.