- PA5-51039 - Invitrogen Antibodies PML antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of PML in 293T-UV 22RV1 cells. Samples were probed using PML Polyclonal Antibody (Product # PA5-51039).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of PML in A549 cells. Samples were probed using PML Polyclonal Antibody (Product # PA5-51039).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockdown of PML was achieved by transfecting HeLa with PML specific siRNAs (Silencer® select Products # s10715). Western blot analysis (Fig. a) was performed using modified whole cell extracts (1% SDS) from the PML knockdown cells (Lane 3), non-specific scrambled siRNA transfected cells (Lane 2) and untransfected cells (Lane 1). The blot was probed with PML Polyclonal Antibody (Product # PA5-51039, 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). Reduction of signal upon siRNA mediated knock down confirms that antibody is specific to PML.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot was performed using Anti-PML Polyclonal Antibody (Product # PA5-51039) and a 65 kDa band corresponding to PML was observed in all tested cell lines lysates, and was upregulated upon IFNa treatment in HeLa. Modified whole cell lysates (1% SDS) (30 µg lysate) of HeLa (Lane 1), HeLa treated with IFNa (1000U, 8 hours) (Lane 2), A549 (Lane 3) and SK-OV-3 (Lane 4) were electrophoresed using NuPAGE® 10 % Bis-Tris gel (Product # NP0302BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blots were 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 SuperSignal™ West Dura Extended Duration Substrate (Product # 34076).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of PML was performed using HeLa and HeLa cells treated with IFN alpha (1000U for 8hrs). 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 PML Polyclonal Antibody (Product # PA5-51039) at 5 µg/mL 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 (Product # A27034) 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 untreated cells, showing lesser expression of PML. Panel f represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

PA5-51039