PA5-28556

antibody from Invitrogen Antibodies

Targeting: GTF2B TFIIB

Antibody data

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Validation data

Product number: PA5-28556 - Provider product page
Provider: Invitrogen Antibodies
Product name: GTF2B Polyclonal Antibody
Antibody type: Polyclonal
Antigen: Recombinant protein fragment
Description: Recommended positive controls: 293T, A431, HeLa, HepG2, HeLa nuclear extract, GTF2B-transfected 293T. Predicted reactivity: Mouse (99%), Rat (99%), Zebrafish (95%), Xenopus laevis (96%), Rhesus Monkey (100%), Bovine (99%). Store product as a concentrated solution. Centrifuge briefly prior to opening the vial.
Reactivity: Human, Mouse
Host: Rabbit
Isotype: IgG
Vial size: 100 µL
Concentration: 1 mg/mL
Storage: Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.

GTF2B protein structure - PA5-28556 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockdown of GTF2B was achieved by transfecting HeLa cells with GTF2B specific siRNAs (Silencer® select Product # s6295, s6296). Western blot analysis (Fig. a) was performed using modified whole cell extracts (1% SDS) from the GTF2B knockdown cells (Lane 3), non-specific scrambled siRNA transfected cells (Lane 2) and untransfected cells (Lane 1). The blots were probed with Anti-GTF2B Polyclonal Antibody (Product # PA5-28556, 1:1000 dilution) and Goat anti-Rabbit IgG (Heavy Chain), Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4000 dilution). Densitometric analysis of this Western Blot is shown in histogram (Fig. b). Loss of signal upon siRNA mediated knock down confirms that antibody is specific to GTF2B. An uncharacterized band around 110 kDa was also observed.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot was performed using Anti-GTF2B Polyclonal Antibody (Product # PA5-28556) and 36 kDa band corresponding to GTF2B was observed across the cell lines and tissues tested. An uncharacterized band of ~ 110 kDa was also observed in all the cell lines. Modified whole cell extracts (1% SDS) (30 µg lysate) of HeLa (Lane1), K-562 (Lane 2), MOLT-4 (Lane 3), Hep G2 (Lane 4), Jurkat (Lane 5), MCF7 (Lane 6), tissue extracts (30 µg lysate) of Mouse Lungs (Lane 7) and Mouse Spleen (Lane 8) were electrophoresed using Novex® NuPAGE™ 10% Bis-Tris Protein Gel (Product # NP0302BOX). 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 (1:1000 dilution) and detected by chemiluminescence with Goat anti-Rabbit IgG (Heavy Chain), Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot analysis of GTF2B was performed by separating 30 µg of non-transfected (–) and transfected (+) 293T whole cell extracts by 12% SDS-PAGE. Proteins were transferred to a membrane and probed with a GTF2B Polyclonal Antibody (Product # PA5-28556) at a dilution of 1:2000. The HRP-conjugated anti-rabbit IgG antibody was used to detect the primary antibody.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot analysis of GTF2B was performed by separating 30 µg of various whole cell extracts by 12% SDS-PAGE. Proteins were transferred to a membrane and probed with a GTF2B Polyclonal Antibody (Product # PA5-28556) at a dilution of 1:1000 and a HRP-conjugated anti-rabbit IgG secondary antibody.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunocytochemistry-Immunofluorescence analysis of GTF2B was performed in HeLa cells fixed in 4% paraformaldehyde at RT for 15 min. Green: GTF2B Polyclonal Antibody (Product # PA5-28556) diluted at 1:1000. Red: phalloidin, a cytoskeleton marker. Scale bar = 10 µm.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of GTF2B 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 1 hour at room temperature. The cells were labeled with GTF2B Polyclonal Antibody (Product # PA5-28556) at 1:100 dilution in 0.1% BSA, incubated at 4 degree Celsius overnight and then with Goat anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32731) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: Green). Nuclei (Panel b: Blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: Red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing nuclear localization. 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: Immunocytochemistry-Immunofluorescence analysis of GTF2B was performed in HeLa cells fixed in 4% paraformaldehyde at RT for 15 min. Green: GTF2B Polyclonal Antibody (Product # PA5-28556) diluted at 1:1000. Red: phalloidin, a cytoskeleton marker. Scale bar = 10 µm.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of GTF2B 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 1 hour at room temperature. The cells were labeled with GTF2B Polyclonal Antibody (Product # PA5-28556) at 1:100 dilution in 0.1% BSA, incubated at 4 degree Celsius overnight and then with Goat anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32731) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: Green). Nuclei (Panel b: Blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: Red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing nuclear localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.