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Western blot
ImmunoprecipitationAntibody data
- Antibody Data
- Antigen structure
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- Validations
- Western blot [2]
- Immunocytochemistry [1]
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Validation data
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- Product number
- PA1-12393 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- PP2A alpha Polyclonal Antibody
- Antibody type
- Polyclonal
- Antigen
- Synthetic peptide
- Reactivity
- Human, Mouse, Rat, Bovine
- Host
- Sheep
- Isotype
- IgG
- Vial size
- 100 µg
- Storage
- -20° C, Avoid Freeze/Thaw Cycles
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Knockdown of PP2A alpha was achieved by transfecting PC-3 cells with PP2A alpha specific siRNAs (Silencer® select Product # s10958). Western blot analysis (Fig. a) was performed using whole cell extracts from the PP2A alpha knockdown cells (Lane 3), non-specific scrambled siRNA transfected cells (Lane 2) and untransfected cells (Lane 1). The blot was probed with Anti-PP2A alpha Polyclonal Antibody (Product # PA1-12393, 1:1000 dilution) and Rabbit anti-Sheep IgG (H+L) Secondary Antibody, HRP (Product # 61-8620, 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 PP2A alpha.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-PP2A alpha Polyclonal Antibody (Product # PA1-12393) and 36 kDa band corresponding to PP2A alpha was observed across the cell lines and tissues tested. Whole cell extracts (30 µg lysate) of HL-60 (Lane 1), A-431 (Lane2), PC-3 (Lane 3), HEL 92.1.7 (Lane 4), K-562 (Lane 5), A549 (Lane 6), HeLa (Lane 7), tissue extracts (30 µg lysate) of Mouse Brain (Lane 8) and Mouse Kidney (Lane 9) 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 Rabbit anti-Sheep IgG (H+L) Secondary Antibody, HRP (Product # 61-8620, 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 PP2A alpha was performed using 70% confluent log phase PC-3 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 PP2A alpha Polyclonal Antibody (Product # PA1-12393) at 1:100 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then with Donkey anti-Sheep IgG (H+L) Cross-Adsorbed Secondary Antibody, Alexa Fluor 488 (Product # A-11015) at a dilution of 1:2000 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). Panel d represents the merged image showing nuclear, cytoskeletal and cytoplasmic localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.
