- PA5-30216 - Invitrogen Antibodies EIF4A1 antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western Blot using eIF4A1 Polyclonal Antibody (Product # PA5-30216). Sample (30 µg of whole cell lysate). Lane A: HeLa. 10% SDS PAGE. eIF4A1 Polyclonal Antibody (Product # PA5-30216) diluted at 1:1,000. 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 eIF4A1 was achieved by transfecting HeLa with eIF4A1 specific siRNAs (Silencer® select Product # s4567, s4568). Western blot analysis (Fig. a) was performed using Whole cell extracts from the eIF4A1 knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with eIF4A1 Polyclonal Antibody (Product # PA5-30216, 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 eIF4A1.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot was performed using Anti-eIF4A1 Polyclonal Antibody(Product # PA5-30216) and a 46kDa band corresponding to eIF4A1 was observed across the cell lines tested. Whole cell extracts (30 µg lysate) of MCF7 (Lane 1), Hep G2 (Lane 2), HeLa (Lane 3), A-431 (Lane 4) and A549 (Lane 5) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). Resolved proteins were then transferred onto a Nitrocellulose membrane (Product # LC2001) 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 EIF4A1 in methanol-fixed HeLa cells using an EIF4A1 polyclonal antibody (Product # PA5-30216) at a 1:200 dilution.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence analysis of eIF4A1 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 10 minutes, and blocked with 2% BSA for 45 minutes at room temperature. The cells were labeled with eIF4A1 Polyclonal Antibody (Product # PA5-30216) at 1:200 dilution 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 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 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: eIF4A1 Polyclonal Antibody detects EIF4A1 protein at cytosol on human colon carcinoma by immunohistochemical analysis. Sample: Paraffin-embedded human colon carcinoma. EIF4A1 Polyclonal Antibody (Product # PA5-30216) dilution: 1:500. Antigen Retrieval: EDTA based buffer, pH 8.0, 15 min.

PA5-30216