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Western blotAntibody data
- Antibody Data
- Antigen structure
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- Validations
- Western blot [3]
- Immunocytochemistry [2]
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- Product number
- PA5-20974 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- FOXO4 Polyclonal Antibody
- Antibody type
- Polyclonal
- Antigen
- Synthetic peptide
- Description
- A suggested positive control is Hela cell lysate. PA5-20974 can be used with blocking peptide PEP-1088.
- Reactivity
- Human, Mouse, Rat
- Host
- Rabbit
- Isotype
- IgG
- Vial size
- 100 µg
- Concentration
- 1 mg/mL
- Storage
- Maintain refrigerated at 2-8°C for up to 3 months. For long term storage store at -20°C
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western Blot analysis of FOXO4 in HeLa cell lysate with FOXO4 Polyclonal Antibody (Product # PA5-20974) at (A) 0.5 and (B) 1 µg/mL.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Knockdown of FOXO4 was achieved by transfecting BeWo with FOXO4 specific siRNAs (Silencer® select Product # S8837, S8835). Western blot analysis (Fig. a) was performed using Whole cell extracts from the FOXO4 knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with FOXO4 Polyclonal Antibody (Product # PA5-20974, 0.5 µg/mL ) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4000). Densitometric analysis of this western blot is shown in histogram (Fig. b). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to FOXO4.Both isoforms showed knockdown compared to scrambled and un-transfected lysates. 53kDa was knocked down completely.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-FOXO4 Polyclonal Antibody (Product # PA5-20974) and a 47 and 53 kDa band corresponding to FOXO4 was observed across cell lines and tissues tested . Whole cell extracts (30 µg lysate) of Jurkat (Lane 1), BeWo (Lane 2), HEK-293 (Lane 3), Hep G2 (Lane 4), HeLa (Lane 5), RAW 264.7 (Lane 6), NIH/3T3 (Lane 7), PC-12 (Lane 8), Mouse Liver (Lane 9), Rat Liver (Lane 10), Mouse Placenta (Lane 11) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). Resolved proteins were then transferred onto a Nitrocellulose membrane (Product # LC2001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1 µg/mL) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4000) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Reagent Kit (Product # WP20005).Both isoforms showed pick up in BeWo and PC-12 cell lines as seen.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunocytochemistry of FOXO4 in HeLa cells with FOXO4 Polyclonal Antibody (Product # PA5-20974) at 10 µg/mL.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence analysis of FOXO4 was performed using 70% confluent log phase HeLa cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 2% BSA for 45 minutes at room temperature. The cells were labeled with FOXO4 Polyclonal Antibody (Product # PA5-20974) at 10 µg/mL in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Donkey anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32790), (1:2000), for 45 minutes at room temperature (Panel a: Blue). Nuclei (Panel b:Green) were stained with ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). F-actin (Panel c: Red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing Cytosolic localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.
