- PA5-19947 - Invitrogen Antibodies AIFM1 antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of K562 cell lysate using a AIF polyclonal antibody (Product # PA5-19947) at 1 µg/mL.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western Blot Validation in Human Cell Lines. Loading: 15 µg of lysates per lane. Antibodies: AIF Polyclonal Antibody (Product # PA5-19947) (1 µg/mL), 1h incubation at RT in 0.05 NFDM/TBST. Secondary: Goat anti-rabbit IgG HRP conjugate at 1:10,000 dilution.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of AIF was achieved by transfecting HeLa with AIF specific siRNAs (Silencer® select Product # S17442, S17440). Western blot analysis (Fig. a) was performed using Whole cell extracts from the AIF untransfected cells (lane 1), non-targeting scrambled siRNA transfected cells (lane 2) and knockdown cells (lane 3). The blot was probed with AIF Polyclonal Antibody (Product # PA5-19947, 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 AIF.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot was performed using Anti-AIF Polyclonal Antibody(Product # PA5-19947) and a 60kDa band corresponding to AIF was observed across cell lines tested. Whole cell extracts (30 µg lysate) of K-562 (Lane 1), Hep G2 (Lane 2), MCF7 (Lane 3), HeLa (Lane 4) 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 (1:1000 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 K562 cells using a AIF polyclonal antibody (Product # PA5-19947) at a 20 µg/mL dilution.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescent analysis of 4% paraformaldehyde-fixed K562 Cells labeling AIF with AIF Polyclonal Antibody (Product # PA5-19947) at 20 µg/mL, followed by goat anti-rabbit IgG secondary antibody at 1:500 dilution (green).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence analysis of AIF 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 45 minutes at room temperature. The cells were labeled with AIF Polyclonal Antibody (Product # PA5-19947) at 5 µg in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034), (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 dilution). Panel d represents the merged image showing mitochondrial 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)
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Experimental details: Immunocytochemistry staining of Jurkat cells using a AIF polyclonal antibody (Product # PA5-19947) at a 2 µg/mL dilution.

PA5-19947