- MA1-26745 - Invitrogen Antibodies HNRNPL antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of HNRNPL was achieved by transfecting HeLa with HNRNPL specific siRNAs (Silencer® select Product # s6740, s6741). Western blot analysis (Fig. a) was performed using modified whole cell extracts (1% SDS) from the HNRNPL knockdown cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with hnRNP L Monoclonal Antibody (4D11) (Product # MA1-26745, 1:1000 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 and Fig c). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to HNRPL.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot was performed using Anti-hnRNP L Monoclonal Antibody (4D11) (Product # MA1-26745) and 64 kDa band along with a 58 kDa band, which are isoforms corresponding to HNRNPL was observed across cell lines and tissue extracts tested except Mouse Heart which is reported to be low. Modified whole cell extracts (1% SDS) (30 µg lysate) of HeLa (Lane 1), Hep G2 (Lane 2), MCF-7 (Lane 3), Jurkat (Lane 4), NIH/3T3 (Lane 5) and tissue extracts of Mouse Heart (Lane 6), Mouse Thymus (Lane 7) and Mouse Lung (Lane 8) were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris 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-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)
Main image
Experimental details: Immunofluorescence analysis of hnRNP L 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 1 hour at room temperature. The cells were labeled with hnRNP L Mouse Monoclonal Antibody (4D11) (Product # MA1-26745) at 5 µg/mL in 0.1% BSA, incubated at 4 degree celsius overnight and then with Goat anti-Mouse IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32723) at a dilution of 1:2000 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). Panel d represents the merged image showing nuclear localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

MA1-26745