- MA5-25050 - Invitrogen Antibodies IFIT1 antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of IFIT1 in HEK293T cells in untransfected (Left lane) and transfected (Right lane) samples using 5 µg per lane. The samples were separated by SDS-PAGE and probed with IFIT1 (Product # MA5-25050) monoclonal antibody.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockout of IFIT1 was achieved by CRISPR-Cas9 genome editing using LentiArray™ Lentiviral sgRNA (Product # A32042, Assay ID CRISPR727742_LV) and LentiArray Cas9 Lentivirus (Product # A32064). Western blot analysis of IFIT1 was performed by loading 50 µg of HeLa Cas9 (Lane 1), HeLa Cas9 treated with 100U/mL IFN beta for 20 hrs (Lane 2) ,HeLa IFIT1 KO (Lane 3) and HeLa IFIT1 KO treated with 100U/mL IFN beta for 20 hrs (Lane 4) whole cell extracts. The samples were electrophoresed using NuPAGE™ Novex™ 4-12% Bis-Tris Protein Gel (Product # NP0322BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with Anti-IFIT1 Monoclonal Antibody (OTI3G8) (Product # MA5-25050, 1:500 dilution) and Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:10,000 dilution) using the iBright™ FL 1500 (Product # A44115). Chemiluminescent detection was performed using SuperSignal™ West Dura Extended Duration Substrate (Product # 34076). Loss of signal upon CRISPR mediated knockout (KO) using the LentiArray™ CRISPR product line confirms that antibody is specific to IFIT1.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of IFIT1 in human tissue (1: Testis; 2: Omentum; 3: Uterus; 4: Breast; 5: Brain; 6: Liver; 7: Ovary; 8: Thyroid gland; 9: colon;10: spleen) samples using 10 µg per lane. Samples were separated by SDS-PAGE and probed with IFIT1 (Product # MA5-25050) monoclonal antibody at a dilution of 1:200.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of Interferon-induced protein with tetratricopeptide repeats 1 was achieved by transfecting HeLa treated with Human IFN Beta (1000 U/mL for 24hrs) with Interferon-induced protein with tetratricopeptide repeats 1 specific siRNAs (Silencer® select Product # s7150, s7151). For detectable levels of IFIT1, transfected cells were treated with Human IFN Beta. Western blot analysis (Fig. a) was performed using Whole cell extracts from the Interferon-induced protein with tetratricopeptide repeats 1 knockdown cells treated with Human IFN Beta (lane 6), IFIT3 control HeLa cells (Lane 5), non-targeting scrambled siRNA transfected HeLa cells treated with Human IFN Beta (lane 4), non-targeting scrambled siRNA transfected HeLa cells (lane 3), Untransfected HeLa cells treated with Human IFN Beta (Lane 2) and untransfected HeLa cells (lane 1) The blot was probed with IFIT1 Monoclonal Antibody (OTI3G8) (Product # MA5-25050, 1:500 dilution) and Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 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 1.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot was performed using Anti-IFIT1 Monoclonal Antibody (OTI3G8) (Product # MA5-25050) and a 55 kDa band corresponding to Interferon-induced protein with tetratricopeptide repeats 1 was observed across cell lines tested. Whole cell extracts (30 µg lysate) of A549 (Lane 1), A549 treated with Human IFN Beta (1000U/mL for 24 hrs) (Lane 2), HT-1080 (Lane 3), HT-1080 treated with Human IFN Beta (1000U/mL for 24 hrs) (Lane 4), HeLa (Lane 5), HeLa treated with Human IFN Beta (1000U/mL for 24 hrs) (Lane 6) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). 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 (1:500 dilution) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 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 IFIT1 in COS7 cells. Cells were transfected with a plasmid overexpressing IFIT1 and probed with a IFIT1 monoclonal antibody (Product # MA5-25050).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescent analysis of IFIT1 in COS7 cells. Cells were transfected with a plasmid overexpressing IFIT1 and probed with a IFIT1 monoclonal antibody (Product # MA5-25050).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence analysis of Interferon-induced protein with tetratricopeptide repeats 1 was performed using 70% confluent log phase HeLa treated with Human IFN Beta (1000 U/mL for 16 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 IFIT1 Monoclonal Antibody (OTI3G8) (Product # MA5-25050) at 1:100 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Donkey anti-Mouse IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32766), (1:2,000 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 in EVOS™ M7000 Imaging System (Product # AMF7000) and externally deconvoluted (D.Sage et al. / Methods 115 (2017) 28–41).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemistry was performed on paraffin-embedded human liver tissue. To expose target proteins, 10mM citric buffer, pH6.0, 100°C for 10min was used. Following antigen retrieval, tissues were probed with a IFIT1 monoclonal antibody (Product # MA5-25050).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemistry was performed on paraffin-embedded adenocarcinoma of human ovary tissue. To expose target proteins, 10mM citric buffer, pH6.0, 100°C for 10min was used. Following antigen retrieval, tissues were probed with a IFIT1 monoclonal antibody (Product # MA5-25050).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemistry was performed on paraffin-embedded carcinoma of human liver tissue. To expose target proteins, 10mM citric buffer, pH6.0, 100°C for 10min was used. Following antigen retrieval, tissues were probed with a IFIT1 monoclonal antibody (Product # MA5-25050).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemistry was performed on paraffin-embedded human kidney tissue. To expose target proteins, 10mM citric buffer, pH6.0, 100°C for 10min was used. Following antigen retrieval, tissues were probed with a IFIT1 monoclonal antibody (Product # MA5-25050).

MA5-25050

Antibody data