- PA5-28160 - Invitrogen Antibodies PRKACA antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of PKA alpha using 30 µg of HeLa lysate. Samples were loaded onto a 10% SDS-PAGE gel and probed with a PKA alpha polyclonal antibody (Product # PA5-28160) at a dilution of 1:1000.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot using PKA alpha Polyclonal Antibody (Product # PA5-28160). Sample (50 µg of whole cell lysate). Lane A: Mouse brain. 10% SDS PAGE. PKA alpha Polyclonal Antibody (Product # PA5-28160) diluted at 1:1,000.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of PKA C-alpha was achieved by transfecting NIH:OVCAR-3 cells with PKA C-alpha specific siRNAs (Silencer® select Product # s11605, s11607). Western blot analysis (Fig. a) was performed using whole cell extracts from Protein kinase A knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with PKA alpha Polyclonal Antibody (Product # PA5-28160, 1:1000 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:10,000 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 PKA C-alpha.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot using PKA alpha Polyclonal Antibody (Product # PA5-28160). Various whole cell extracts (30 µg) were separated by 10% SDS-PAGE, and the membrane was blotted with PKA alpha Polyclonal Antibody (Product # PA5-28160) 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: Western blot was performed using PKA C-alpha Polyclonal Antibody (Product # PA5-28160) and a 41 kDa band corresponding to PKA C-alpha along with a non-specific band at ~ 60 kDa was observed across cell lines and tissues tested. Whole cell extracts (30 µg lysate) of A549 (Lane 1), HeLa (Lane 2), Jurkat (Lane 3), Hep G2 (Lane 4), and tissue extracts (30 µg lysate) of Mouse Brain (Lane 5), Rat Brain (Lane 6) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX), 10 well. 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, 1:10,000 dilution) using the iBright™ FL1500 Imaging System (Product # A44115). Chemiluminescentdetection 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 PKA alpha in methanol-fixed HeLa cells using a PKA alpha polyclonal antibody (Product # PA5-28160) at a 1:200 dilution.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence analysis of PKA C-alpha was performed using 70% confluent log phase NIH:OVCAR-3 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 PKA alpha Polyclonal Antibody (Product # PA5-28160) at 1:200 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Donkey anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor™ Plus 488 (Product # A32790, 1:2500 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). Panel d represents the merged image showing cytosolic 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: Immunohistochemical analysis of paraffin-embedded human colon, using PKA alpha (Product # PA5-28160) antibody at 1:100 dilution. Antigen Retrieval: EDTA based buffer, pH 8.0, 15 min.

PA5-28160