- PA5-27361 - Invitrogen Antibodies LYN antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Lyn Polyclonal Antibody detects LYN protein by Western blot analysis. A. 30 µg Raji whole cell lysate/extract. B. 30 µg K562 whole cell lysate/extract. C. 30 µg Raji whole cell lysate/extract.7.5 % SDS-PAGE. Lyn Polyclonal Antibody (Product # PA5-27361) dilution: 1:1,000.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western Blot using Lyn Polyclonal Antibody (Product # PA5-27361). Mouse tissue extract (50 µg) was separated by 10% SDS-PAGE, and the membrane was blotted with Lyn Polyclonal Antibody (Product # PA5-27361) diluted at 1:500. 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: Knockdown of Tyrosine-protein kinase Lyn was achieved by transfecting K-562 with Tyrosine-protein kinase Lyn specific siRNAs (Silencer® select Product # S8356, S8355). Western Blot analysis (Fig. a) was performed using Whole cell extracts from the Tyrosine-protein kinase Lyn knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The Blot was probed with Lyn Polyclonal Antibody (Product # PA5-27361, 1:1000 ) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4000). 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 Tyrosine-protein kinase Lyn.^^13

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western Blot was performed using Anti-Lyn Polyclonal Antibody (Product # PA5-27361) and a 58 kDa band corresponding to Tyrosine-protein kinase Lyn was observed across cell lines and tissues. Whole cell extracts (30 µg lysate) of K-562 (Lane 1), Raji (Lane 2), A549 (Lane 3), Jurkat (Lane 4), Mouse Spleen (Lane 5), Rat Spleen (Lane 6), Mouse Brain (Lane 7), Rat Brain (Lane 8) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein 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) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4000) 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 LYN in paraformaldehyde-fixed A431 cells using a LYN polyclonal antibody (Product # PA5-27361) (Green) at a 1:500 dilution. Alpha-tubulin filaments were labeled with Product # PA5-29281 (Red) at a 1:2000.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence analysis of Lyn 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 Lyn Polyclonal Antibody (Product # PA5-27361) at 1:100 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then with Donkey anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32790) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with Hoechst 33342 (Product # H1399). F-actin (Panel c: red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing cytoplasmic localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 40X magnification in CellInsight CX7 LZR High-Content Screening (HCS) Platform (Product # CX7C1115LZR).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical analysis of paraffin-embedded human colon carcinoma, using LYN (Product # PA5-27361) antibody at 1:250 dilution. Antigen Retrieval: EDTA based buffer, pH 8.0, 15 min.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Lyn Polyclonal Antibody immunoprecipitates LYN protein in IP experiments. IP samples: K562 whole cell extract. A. Control with 4 µg of preimmune Rabbit IgG. B. Immunoprecipitation of LYN protein by 4 µg Lyn Polyclonal Antibody (Product # PA5-27361). 5 % SDS-PAGE. The immunoprecipitated LYN protein was detected by Lyn Polyclonal Antibody (Product # PA5-27361) diluted at 1:500.

PA5-27361