- PA5-29489 - Invitrogen Antibodies DYNC1I1 antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western Blot using DYNC1I1 Polyclonal Antibody (Product # PA5-29489). Sample (30 µg of whole cell lysate). Lane A: Raji. Lane B: K562. Lane C: HL-60 . 7.5% SDS PAGE. DYNC1I1 Polyclonal Antibody (Product # PA5-29489) diluted at 1:1,000.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: KD of DYNC1I1 was achieved by transfecting U-87 MG with DYNC1I1 specific siRNAs (Silencer® select Product # s4203, s4204). Western blot analysis (Fig. a) was performed using whole cell extracts from the DYNC1I1 KD cells (Lane 3), non-specific scrambled siRNA transfected cells (Lane 2) and untransfected cells (Lane 1). The blot was probed with DYNC1I1 Polyclonal Antibody (Product # PA5-29489, 1:1500 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.25µg/mL, 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 DYNC1I1..

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot was performed using Anti-DYNC1I1 Polyclonal Antibody (Product # PA5-29489) and a 70 kDa band corresponding to DYNC1I1 was observed across cell lines and tissues tested. Whole cell extracts (30 µg lysate) of K-562 (Lane 1), Raji (Lane 2), Jurkat (Lane 3), U-87 MG (Lane 4), SH-SY5Y (Lane 5), SK-OV-3 (Lane 6), Rat Brain (Lane 7) and Mouse Brain (Lane 8) were electrophoresed using NuPAGE® 4-12 % Bis-Tris gel (Product # NP0321BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # LC2001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1:1500 dilution) and detected by chemiluminescence Goat Anti-Rabbit IgG Secondary Antibody, HRP conjugate (Product # A27036, 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)
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Experimental details: Immunofluorescence analysis of DYNC1I1 was performed using 70% confluent log phase U-87 MG 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 DYNC1I1 Polyclonal Antibody (Product # PA5-29489) at 1:100 dilution in 0.1% BSA, incubated at 4 degree Celsius overnight and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (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 cytoskeleton localization. Panel e represents 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 U87 xenograft, using DYNC1I1 (Product # PA5-29489) antibody at 1:500 dilution. Antigen Retrieval: EDTA based buffer, pH 8.0, 15 min.

PA5-29489