720128

antibody from Invitrogen Antibodies

Targeting: FOXO3 AF6q21, FKHRL1, FOXO2, FOXO3A

Western blot (Supportive data in Antibodypedia)

Immunocytochemistry (Supportive data in Antibodypedia)

Flow cytometry (Supportive data in Antibodypedia)

Chromatin Immunoprecipitation (Supportive data in Antibodypedia)

Other assay (Supportive data in Antibodypedia)

Antibody Data

Product number

720128

Provider

Invitrogen Antibodies

Product name

FOXO3A Polyclonal Antibody

Antibody type

Polyclonal

Antigen

Synthetic peptide

Reactivity

Human, Mouse

Host

Rabbit

Isotype

IgG

Vial size

100 µg

Concentration

0.5 mg/mL

Storage

Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.

References

Submitted references

FoxO3a Inhibits Tamoxifen-Resistant Breast Cancer Progression by Inducing Integrin α5 Expression.

Ricci E, Fava M, Rizza P, Pellegrino M, Bonofiglio D, Casaburi I, Lanzino M, Giordano C, Bruno R, Sirianni R, Barone I, Sisci D, Morelli C
Cancers 2022 Jan 2;14(1)

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The neuroprotective mechanism of lithium after ischaemic stroke.

Chen B, Zhang M, Ji M, Zhang D, Chen B, Gong W, Li X, Zhou Y, Dong C, Wen G, Zhan X, Wu X, Yuan H, Xu E, Xia M, Verkhratsky A, Li B
Communications biology 2022 Feb 3;5(1):105

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Etv6 activates vegfa expression through positive and negative transcriptional regulatory networks in Xenopus embryos.

Li L, Rispoli R, Patient R, Ciau-Uitz A, Porcher C
Nature communications 2019 Mar 6;10(1):1083

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis was performed on whole cell extracts (50 µg lysate) of HEL (Lane1), Mouse Placenta (Lane 2) and Mouse Liver (Lane 3). The blots were probed with Anti-FOXO3A Rabbit Polyclonal Antibody (Product # 720128, 1-2 µg/mL) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.4 µg/mL, 1:2500 dilution). A 71 kDa band corresponding FOXO3A was observed in cell line and tissues tested. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 4-12% Bis-Tris gel (Product # NP0321BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody following blocking with 5% skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).

Immunocytochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunofluorescence was performed on fixed and permeabilized HeLa cells for detection of FOXO3A using Anti-FOXO3A Rabbit Polyclonal Antibody (Product # 720128, 2 µg/mL) and labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034, 1:2000). Panel a) shows representative cells that were stained for detection and localization of FOXO3A protein (green), Panel b) is stained for nuclei (blue) using SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). Panel c) represents cytoskeletal F-actin staining using Alexa Fluor® 555 Rhodamine Phalloidin (Product # R415, 1:300). Panel d) is a composite image of Panels a, b and c clearly demonstrating nuclear localization of FOXO3A. Panel e) represents control cells with no primary Antibody to assess background.

Flow cytometry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Flow Cytometry analysis of FOXO3A was performed on Hela cells labeled with Anti-FOXO3A Rabbit Polyclonal Antibody (Product# 720128, 2- 4 ug/ 1M cells) or with rabbit isotype control and detected with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, (Alexa Fluor® 488 conjugate, Product # A27034, 0.4 ug/ml, 1:2500) as represented by the red and yellow histograms respectively. The purple histogram represents unstained control cells and the green histogram represents no-primary-antibody control. A representative of 10,000 cells were acquired and analyzed for each sample using an Attune® Acoustic Focusing Cytometer (4468770).

Chromatin Immunoprecipitation

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Chromatin Immunoprecipitation (ChIP) was performed using Anti-FOXO3a Rabbit Polyclonal Antibody (Product # 720128, 3 µg) on sheared chromatin from 2 million Jurkat cells serum starved for 24 hours using the MAGnify ChIP system kit (Product # 49-2024). Normal Rabbit IgG was used as a negative IP control. The purified DNA was analyzed by 7500 Fast qPCR system (Product # 4351106) with optimized PCR primer pairs for the promoters of the active BCL10 region used as positive control target gene, and the region of the inactive SAT2 satellite repeat, used as negative control target gene. Data is presented as fold enrichment of the antibody signal versus the negative control IgG using the comparative CT method.

Other assay

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Fig. 2 Identification of Etv6 direct transcriptional targets in the somites. a Experimental design. RNA-seq was performed on wild type (WT) and Etv6-deficient ( etv6 MO-injected embryos) somite explants dissected from stage 22 embryos. Differentially expressed genes (DEGs) were identified by comparing the transcriptome of these tissues. b Spearman correlation analysis on triple biological RNA-seq replicates. c MA plot (magnitude of the difference versus amplitude of the signal) showing the fold-change in gene expression of all genes in WT versus Etv6-deficient somites (log 2 FC) compared to their expression levels (log 2 CPM). Black dots, non-significant change; red dots, differentially expressed genes (DEGs). The number of positively and negatively regulated DEGs is indicated. d WISH showing the expression of DEGs in stage 22 WT and Etv6-deficient embryos. Meox2 expression is upregulated in the somites (arrows) in Etv6-deficient embryos whereas expression of crim1 , sox18 and vegfa is downregulated. Note that vegfa expression in the hypochord is unaffected (arrowheads). Embryos are shown in lateral view with anterior to the left and dorsal to the top. Numbers in top right corner indicate the number of embryos exhibiting the phenotype pictured (scale bars: 0.5 mm). e The intersection between DEGs and genes harbouring Etv6 ChIP-seq peaks in their TSS region reveals 540 putative direct target genes. f Foxo3 and klf4 (arrows), known transcriptional regul