- PA5-97854 - Invitrogen Antibodies IFIT3 antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western Blot analysis of precipitated IFIT3 from HepG2 whole cell lysate using a IFIT3 polyclonal antibody (Product # PA5-97854). An HRP-conjugated Protein G antibody was used as the secondary antibody (1:2000). Lane 1: Rabbit control IgG (1µg). Lane 2: HepG2 whole cell lysate (500µg). Lane 3: HepG2 whole cell lysate (10µg).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot analysis of precipitated IFIT3 from HepG2 whole cell lysate using a IFIT3 polyclonal antibody (Product # PA5-97854). An HRP-conjugated Protein G antibody was used as the secondary antibody (1:2000). Lane 1: Rabbit control IgG (1µg). Lane 2: HepG2 whole cell lysate (500µg). Lane 3: HepG2 whole cell lysate (10µg).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western Blot analysis of IFIT3 using a IFIT3 Polyclonal antibody (Product # PA5-97854) at a concentration of 3.3 µg/mL. Positive WB detected in: A549 whole cell lysate. A secondary Goat polyclonal antibody to rabbit IgG was applied at a 1:50,000 dilution. Observed band size: 56 kDa.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of Interferon-induced protein with tetratricopeptide repeats 3 was achieved by transfecting A549 cells treated with Human IFN Alpha A (500 U/mL for 24hrs) with Interferon-induced protein with tetratricopeptide repeats 3 specific siRNAs (Silencer® select Product # s7153, s7155). For detectable levels of IFIT3, transfected cells were treated with Human IFN Alpha A. Western blot analysis (Fig. a) was performed using Whole cell extracts from the Interferon-induced protein with tetratricopeptide repeats 3 knockdown cells treated with Human IFN Alpha (lane 6), IFIT3 control A549 cells (Lane 5), non-targeting scrambled siRNA transfected A549 cells treated with Human IFN Alpha A (lane 4), non-targeting scrambled siRNA transfected A549 cells (lane 3), Untransfected A549 cells treated with Human IFN Alpha A (Lane 2) and untransfected A549 cells (lane 1). The blot was probed with IFIT3 Polyclonal Antibody (Product # PA5-97854, 1:1000 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4000 dilution). 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 Interferon-induced protein with tetratricopeptide repeats 3.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot was performed using Anti-IFIT3 Polyclonal Antibody (Product # PA5-97854) and a 55 kDa band corresponding to Interferon-induced protein with tetratricopeptide repeats 3 was observed across cell lines tested. Whole cell extracts (30 µg lysate) of A549 (Lane 1), A549 treated with IFN Alpha A (500 U/mL for 24hrs) (Lane 2), Daudi (Lane 3), Daudi treated with Human IFN Alpha A (0.1 nm for 24hrs) (Lane 4) were electrophoresed using 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 (1:1000 dilution) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L) 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)
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Experimental details: Immunofluorescent analysis of IFIT3 in HepG2 cells using a IFIT3 polyclonal antibody (Product # PA5-97854) at a dilution of 1:100. Alexa Fluor 488-congugated Goat Anti-Rabbit IgG(H+L) secondary antibody was used.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence analysis of Interferon-induced protein with tetratricopeptide repeats 3 was performed using 70% confluent log phase A549 treated with Human IFN Alpha (500 U/mL for 24 hrs). 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 IFIT3 Polyclonal Antibody (Product # PA5-97854) at 1:100 dilution 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 dilution), for 45 minutes at room temperature (Panel a: Green). Nuclei (Panel b:Blue) 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 dilution). Panel d represents the merged image showing cytoplasmic localization. Panel e represents no signal in untreated cells Panel f represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical analysis of IFIT3 in paraffin embedded human colon cancer using a IFIT3 polyclonal antibody (Product # PA5-97854) at a dilution of 1:100.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemical analysis of IFIT3 in paraffin embedded human kidney tissue using a IFIT3 polyclonal antibody (Product # PA5-97854) at a dilution of 1:100.

PA5-97854