- PA5-28344 - Invitrogen Antibodies RAD21 antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of RAD21 using Various whole cell extracts (30 µg). Samples were loaded onto a 7.5% SDS-PAGE gel and probed with a RAD21 polyclonal antibody (Product # PA5-28344) at a dilution of 1:1000.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot analysis of RAD21 was performed by separating 30 µg of various whole cell extracts by 5% SDS-PAGE. Proteins were transferred to a membrane and probed with a RAD21 Polyclonal Antibody (Product # PA5-28344) at a dilution of 1:1000 and a HRP-conjugated anti-rabbit IgG secondary antibody.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot analysis of RAD21 was performed by separating 30 µg of various whole cell extracts by 5% SDS-PAGE. Proteins were transferred to a membrane and probed with a RAD21 Polyclonal Antibody (Product # PA5-28344) at a dilution of 1:1000 and a HRP-conjugated anti-rabbit IgG secondary antibody.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western Blot using RAD21 Polyclonal Antibody (Product # PA5-28344). Sample (30 µg of whole cell lysate). Lane A: NIH-3T3. Lane B: JC. Lane C: BCL-1. 7.5% SDS PAGE. RAD21 Polyclonal Antibody (Product # PA5-28344) diluted at 1:3,000. 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 Anti-RAD21 Polyclonal Antibody (Product # PA5-28344) and a 90 kDa band corresponding to RAD21 was observed across tested cell lines. Nuclear enriched extracts (40 µg lysate) of HeLa (Lane 1), THP-1 (Lane 2), A-431 (Lane 3), DU 145 (Lane 4) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). 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 # A27036,1:20000 dilution) using the iBright™ FL1500 Imaging System (Product # A44115). Chemiluminescent detection was performed using SuperSignal™ West Pico PLUS Chemiluminescent Substrate (Product # 34580).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of RAD21 was achieved by transfecting HeLa with RAD21 specific siRNAs (Silencer® select Product # s531211, s531212). Western blot analysis (Fig. a) was performed using Nuclear enriched extracts from the RAD21 knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with RAD21 Polyclonal Antibody (Product # PA5-28344, 1:1000 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:20000 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 RAD21.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot using RAD21 Polyclonal Antibody (Product # PA5-28344) and a band at ~90 kDa corresponding to RAD21 was decreased upon etoposide treatment in HeLa. Modified whole cell extracts (1% SDS) (30 µg lysate) of HeLa (Lane 1), HeLa treated with 50µM Etoposide for 48 hrs (Lane 2), 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 Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (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). Altered expression of proteins upon cell treatment demonstrates antibody specificity.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunocytochemistry-Immunofluorescence analysis of RAD21 was performed in HeLa cells fixed in 4% paraformaldehyde at RT for 15 min. Green: RAD21 Polyclonal Antibody (Product # PA5-28344) diluted at 1:1000. Red: phalloidin, a cytoskeleton marker.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence analysis of RAD21 was performed using 70% confluent log phase HeLa 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 RAD21 Polyclonal Antibody (Product # PA5-28344) at 5 µg/mL in 0.1% BSA, incubated at 4 degree Celsius overnight and then with Goat anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32731) 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 nuclear localization in HeLa cells. 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: Immunohistochemistry (Paraffin) analysis of RAD21 was performed in paraffin-embedded human lung papillary adenocarcinoma tissue using RAD21 Polyclonal Antibody (Product # PA5-28344) at a dilution of 1:250.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemistry (Paraffin) analysis of RAD21 was performed in paraffin-embedded mouse testis tissue using RAD21 Polyclonal Antibody (Product # PA5-28344) at a dilution of 1:500. Antigen Retrieval: Citrate buffer, pH 6.0, 15 min.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemistry (Paraffin) analysis of RAD21 was performed in paraffin-embedded rat kidney tissue using RAD21 Polyclonal Antibody (Product # PA5-28344) at a dilution of 1:500. Antigen Retrieval: Citrate buffer, pH 6.0, 15 min.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical analysis of paraffin-embedded C2C12 xenograft, using RAD21 (Product # PA5-28344) antibody at 1:750 dilution. Antigen Retrieval: EDTA based buffer, pH 8.0, 15 min.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical analysis of paraffin-embedded HBL435 xenograft, using RAD21 (Product # PA5-28344) antibody at 1:750 dilution. Antigen Retrieval: EDTA based buffer, pH 8.0, 15 min.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Chromatin Immunoprecipitation (ChIP) assay of endogenous RAD21 protein using RAD21 Antibody: ChIP was performed using RAD21 Polyclonal Antibody (Product # PA5-28344, 5 µg) on sheared chromatin from MCF7 and MCF7 cells treated with Estradiol (100 nM, 45 minutes) using the MAGnify ChIP System kit (Product # 49-2024). Normal Rabbit IgG was used as a negative IP control. The purified DNA was analyzed by qPCR using primers binding to MYC promotor regions (P1 and P2), ERE2 (active) and MYC1 (-394kb of myc TSS), SAT alpha (Inactive). Data is presented as fold enrichment of the antibody signal versus the negative control IgG using the comparative CT method.

PA5-28344

Antibody data