- MA5-25574 - Invitrogen Antibodies PTPN1 antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of PTPN1 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 PTPN1 (Product # MA5-25574) monoclonal antibody.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of PTPN1 in HeLa wild-type and knockout cells using 10 µg per lane. Samples were separated by SDS-PAGE and probed with PTPN1 (Product # MA5-25574) monoclonal antibody and HSP90AB1 antibody with a dilution of 1:500 as a loading control.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of PTP1B was achieved by transfecting MCF-7 cells with PTP1B specific siRNAs (Silencer® select Product # s11508 ). Western blot analysis (Fig. a) was performed using membrane extracts from the PTP1B knockdown cells (Lane 2) and non-specific scrambled siRNA transfected cells (Lane 1). The blot was probed using PTP1B Monoclonal Antibody (Product # MA5-25574) 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 PTP1B.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot was performed using Anti-PTP1B Monoclonal Antibody (Product # MA5-25574). A ~45kDa band corresponding to PTP1B was observed in Jurkat and MCF-7 cell lines. Membrane enriched extracts (30 µg) of Jurkat (Lane 1) and MCF-7 (Lane 2) were electrophoresed using NuPAGE™ 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 the primary antibody (1:2000 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 PTPN1 in COS7 cells. Cells were transfected with a plasmid overexpressing PTPN1 and probed with a PTPN1 monoclonal antibody (Product # MA5-25574).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence analysis of PTP1B was performed using 70% confluent log phase MCF-7 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 60 minutes at room temperature. The cells were labeled with PTPB1 Monoclonal Antibody (Product # MA5-25574) at 1:100 dilution in 0.1% BSA, incubated at 4 degree Celsius overnight and then labeled with Goat anti-Mouse IgG (H+L), Superclonal™ Recombinant Secondary Antibody, Alexa Fluor 488 (Product # A28175), (1:2000 dilution) 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 cytoplasmic localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

MA5-25574