712913

antibody from Invitrogen Antibodies

Targeting: FTO ALKBH9, KIAA1752, MGC5149

Provider product page for 712913

Western blot

Immunocytochemistry

Other assay

Antibody data

Antibody Data
Antigen structure
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Validations
Western blot [2]
Immunocytochemistry [2]
Other assay [1]

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Product number: 712913 - Provider product page
Provider: Invitrogen Antibodies
Product name: FTO Recombinant Superclonal™ Antibody (2HCLC)
Antibody type: Other
Antigen: Other
Description: This antibody is predicted to react with Mouse, Rat, Bovine, Recombinant rabbit Superclonal™ antibodies are unique offerings from Thermo Fisher Scientific. They are comprised of a selection of multiple different recombinant monoclonal antibodies, providing the best of both worlds - the sensitivity of polyclonal antibodies with the specificity of monoclonal antibodies - all delivered with the consistency only found in a recombinant antibody. While functionally the same as a polyclonal antibody - recognizing multiple epitope sites on the target and producing higher detection sensitivity for low abundance targets - a recombinant rabbit Superclonal™ antibody has a known mixture of light and heavy chains. The exact population can be produced in every lot, circumventing the biological variability typically associated with polyclonal antibody production. Note: Formerly called “Recombinant polyclonal antibody”, this product is now rebranded as “Recombinant Superclonal™ antibody”. The physical product and the performance remain unchanged.
Reactivity: Human
Host: Rabbit
Isotype: IgG
Antibody clone number: 2HCLC
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.

FTO protein structure - 712913 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot was performed using Anti-FTO Rabbit Recombinant Rabbit Superclonal™ Antibody (Product # 712913) and a 58kDa band corresponding to FTO was observed across cell line tested. Whole cell extracts (30 µg lysate) of SH-SY5Y (Lane 1), A549 (Lane 2), and THP-1 (Lane 3) were electrophoresed using Novex® NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1:500 dilution) and detected by chemiluminescence Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036) at a 1:5000 dilution. Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockdown of FTO was achieved by transfecting A549 cells with FTO specific siRNA (Silencer® select Product # S35511, Product # S35512) . Western blot analysis (Fig a) was performed using whole cell extracts (1% SDS) from FTO knockdown cells (Lane 3), non-specific scrambled siRNA transfected cells (Lane 2) and untransfected cells (Lane 1). The blot was probed with Anti-FTO Recombinant Rabbit Superclonal™ Antibody (Product # 712913) at a 1:500 dilution and Goat anti-Rabbit IgG (Heavy Chain) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036) (0.25 µg/mL, 1:4000 dilution). Densitometric analysis of this western blot is shown in the histogram (Fig b). Loss of signal upon siRNA mediated knockdown confirms that antibody is specific to FTO.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: For immunocytochemistry analysis, HeLa cells were fixed and permeabilized for detection of endogenous FTO using Anti-FTO Recombinant Rabbit Polycloclonal Antibody (Product # 712913) at a 1:100 dilution and labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034) at a 1:2000 dilution. Panel a) shows representative cells that were stained for detection and localization of FTO protein (green), Panel b) is stained for nuclei (blue) using ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). Panel c) represents cytoskeletal F-actin staining using Rhodamine Phalloidin (Product # R415) at a 1:300 dilution. Panel d) is a composite image of Panels a, b and c clearly demonstrating cytoplasm and nuclear speckles localization of FTO. Panel e) represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: For immunocytochemistry analysis, HeLa cells were fixed and permeabilized for detection of endogenous FTO using Anti-FTO Recombinant Rabbit Polycloclonal Antibody (Product # 712913) at a 1:100 dilution and labeled with Goat anti-Rabbit IgG (Heavy Chain) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034) at a 1:2000 dilution. Panel a) shows representative cells that were stained for detection and localization of FTO protein (green), Panel b) is stained for nuclei (blue) using ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). Panel c) represents cytoskeletal F-actin staining using Rhodamine Phalloidin (Product # R415) at a 1:300 dilution. Panel d) is a composite image of Panels a, b and c clearly demonstrating cytoplasm and nuclear speckles localization of FTO. Panel e) represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.