- PA5-27495 - Invitrogen Antibodies IFITM1 antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of CD225/IFITM1 using 30 µg of A) A549 and B) HeLa lysate. Samples were loaded onto a 15% SDS-PAGE gel and probed with a CD225/IFITM1 polyclonal antibody (Product # PA5-27495) at a dilution of 1:1000.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot was performed using Anti-IFITM1 Polyclonal Antibody (Product # PA5-27495) and a 15 kDa band corresponding to Interferon-induced transmembrane protein 1 was observed across cell lines tested. Membrane enriched extracts (40 µg lysate) of MCF7 (Lane 1), MCF7 treated with Human IFN Alpha (1000 U/mL for 12hrs) (Lane 2), Caco-2 (Lane 3), Caco2 treated with Human IFN Beta 250 IU/mL 24hrs (Lane 4) and K-562 (Lane 5) were electrophoresed using Novex™ 16% Tricine Protein Gel (Product # EC6695BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23002) 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: Western Blot using IFITM1 Polyclonal Antibody (Product # PA5-27495). Various whole cell extracts (30 µg) were separated by 15% SDS-PAGE, and the membrane was blotted with IFITM1 Polyclonal Antibody (Product # PA5-27495) diluted at 1:500. The HRP-conjugated anti-rabbit IgG antibody was used to detect the primary antibody.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of Interferon-induced transmembrane protein 1 was achieved by transfecting MCF7 treated cells with Human IFN Alpha (1000u/mL for 12hrs) with Interferon-induced transmembrane protein 1 specific siRNAs (Silencer® select Product # S16192, S16193). Western Blot analysis (Fig. a) was performed using membrane enriched extracts from the Interferon-induced transmembrane protein 1 knockdown cells treated with Human IFN Alpha (lane 6), IFIT3 control MCF7 cells (Lane 5), non-targeting scrambled siRNA transfected MCF7 cells treated with Human IFN Alpha A (lane 4), non-targeting scrambled siRNA transfected MCF7cells (lane 3), Untransfected MCF7 cells treated with Human IFN Alpha A (Lane 2) and untransfected MCF7 cells (lane 1).. The blot was probed with IFITM1 Polyclonal Antibody (Product # PA5-27495, 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 transmembrane protein 1.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescent analysis of CD225/IFITM1 in methanol-fixed HeLa cells using a CD225/IFITM1 polyclonal antibody (Product # PA5-27495) at a 1:500 dilution.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence analysis of Interferon-induced transmembrane protein 1 was performed using 70% confluent log phase MCF7 treated with Human IFN Alpha (1000u/mL for 12hrs). 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 IFITM1 Polyclonal Antibody (Product # PA5-27495) 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 as well as membrane like localization. Panel e represents no expression 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: IFITM1 Polyclonal Antibody detects IFITM1 protein at cytosol and membrane on human ovarian carcinoma by immunohistochemical analysis. Sample: Paraffin-embedded human ovarian carcinoma. IFITM1 Polyclonal Antibody (Product # PA5-27495) dilution: 1:500. Antigen Retrieval: EDTA based buffer, pH 8.0, 15 min.

PA5-27495