- PA5-98058 - Invitrogen Antibodies PHF6 antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western Blot analysis of precipitated PHF6 from Jurkat whole cell lysate using a PHF6 polyclonal antibody (Product # PA5-98058). An HRP-conjugated Protein G antibody was used as the secondary antibody (1:2000). Lane 1: Rabbit control IgG. Lane 2: Jurkat whole cell lysate (500µg). Lane 3: Jurkat whole cell lysate (10µg).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western Blot analysis of PHF6 using a PHF6 Polyclonal antibody (Product # PA5-98058) at a concentration of 2.8 µg/mL. Positive WB detected in: Hela whole cell lysate, K562 whole cell lysate, Jurkat whole cell lysate. A secondary Goat polyclonal antibody to rabbit IgG was applied at a 1:50,000 dilution. Observed band size: 42 kDa.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of PHD finger protein 6 was achieved by transfecting Jurkat with PHF6 specific siRNAs (Silencer® select Product # s533661, s38847). Western Blot analysis (Fig. a) was performed using acid extracts from the PHF6 knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with PHF6 Polyclonal Antibody (Product # PA5-98058, 1.75 µg/mL) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:10000 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 PHD finger protein 6.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western Blot was performed using Anti-PHF6 Polyclonal Antibody (Product # PA5-98058) and a 41 kDa band corresponding to PHD finger protein 6 was observed across all cell lines tested except Hep G2, which is reported low for PHF6. Uncharacterised bands (*) at ~95 kDa and 145 kDa were also observed in all cell lines. Acid extracts (40 µg lysate) of Jurkat (Lane 1), Hep G2 (Lane 2), HeLa (Lane 3), K-562 (Lane 4), Mouse Thymus (Lane 5), and Rat Brain (Lane 6) were electrophoresed using NuPAGE™ 10% Bis-Tris Protein Gel (Product # NP0301BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23002) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1.75 µg/mL dilution) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:10000 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 PHF6 in Hela cells using a PHF6 polyclonal antibody (Product # PA5-98058) at a dilution of 1:233. The cells were fixed in 4% formaldehyde, permeabilized using 0.2% Triton X-100 and blocked in 10% normal Goat Serum. The cells were then incubated with the antibody overnight at 4°C. The secondary antibody was Alexa Fluor 488-congugated Goat Anti-Rabbit IgG(H+L). Cells were counter-stained with DAPI.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical analysis of PHF6 in paraffin embedded human testis tissue using a PHF6 polyclonal antibody (Product # PA5-98058) at a dilution of 1:100.

PA5-98058