- 710048 - Invitrogen Antibodies RELA antibody

Submitted references Ghrelin ameliorates chronic obstructive pulmonary disease-associated infllammation and autophagy.

Song B, Yan X, Li R, Zhang H
Biotechnology and applied biochemistry 2021 Apr;68(2):356-365

Plasma-derived extracellular vesicles from Plasmodium vivax patients signal spleen fibroblasts via NF-kB facilitating parasite cytoadherence.

Toda H, Diaz-Varela M, Segui-Barber J, Roobsoong W, Baro B, Garcia-Silva S, Galiano A, Gualdrón-López M, Almeida ACG, Brito MAM, de Melo GC, Aparici-Herraiz I, Castro-Cavadía C, Monteiro WM, Borràs E, Sabidó E, Almeida IC, Chojnacki J, Martinez-Picado J, Calvo M, Armengol P, Carmona-Fonseca J, Yasnot MF, Lauzurica R, Marcilla A, Peinado H, Galinski MR, Lacerda MVG, Sattabongkot J, Fernandez-Becerra C, Del Portillo HA
Nature communications 2020 Jun 2;11(1):2761

Systematic Immunotherapy Target Discovery Using Genome-Scale In Vivo CRISPR Screens in CD8 T Cells.

Dong MB, Wang G, Chow RD, Ye L, Zhu L, Dai X, Park JJ, Kim HR, Errami Y, Guzman CD, Zhou X, Chen KY, Renauer PA, Du Y, Shen J, Lam SZ, Zhou JJ, Lannin DR, Herbst RS, Chen S
Cell 2019 Aug 22;178(5):1189-1204.e23

Codonopis bulleynana Forest ex Diels inhibits autophagy and induces apoptosis of colon cancer cells by activating the NF-κB signaling pathway.

Luan Y, Li Y, Zhu L, Zheng S, Mao D, Chen Z, Cao Y
International journal of molecular medicine 2018 Mar;41(3):1305-1314

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of NF-kappa-B (p65) was performed by loading 20 µg of HeLa (lane1), A549 (lane2), A431 (lane3), K562 (lane4), Jurkat (lane5) HEK-293 (lane6) and MDA-MB-231 (lane7) cell lysates using Novex®NuPAGE®4-12 % Bis-Tris gel (Product # NP0321BOX), XCell SureLock Electrophoresis System (Product # EI0002), Novex® Sharp Pre-Stained Protein Standard (Product # LC5800), and iBlot® Dry Blotting System (Product # IB21001). Proteins were transferred to a nitrocellulose membrane and blocked with 5 % skim milk for 1 hour at room temperature. NF-kappa-B (p65) was detected at ~65 kDa using NF-kappa-B (p65) Mouse Recombinant Rabbit Polyclonal Antibody (Product # 710048) at 0.5-1 µg/mL in 2.5 % skim milk at 4°C overnight on a rocking platform. Goat anti-Rabbit IgG-HRP Secondary Antibody (Product # G-21234) at 1:5000 dilution was used and chemiluminescent detection was performed using Pierce™ ECL Western blotting Substrate (Product # 32106).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of NFkB p65 was achieved by transfecting HeLa with NFkB p65 specific siRNAs (Silencer® select Product # s11916, s11914). Western blot analysis was performed using Nuclear enriched extracts from the NFkB p65 knockdown cells (lane 3), scrambled siRNA transfected cells (lane 2), and untransfected cells (lane 1). The blot was probed with NFkB p65 Recombinant Polyclonal Antibody (4-2HCLC) (Product # 710048, 0.5 µg/mL) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:20000 dilution). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to NFkB p65.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of NFkB p65 in HeLa cell lysate (30 µg/lane) using a NFkB p65 Recombinant Rabbit Polyclonal Antibody (Product # 710048) at a dilution of 1 µg/mL. Samples were blocked in 5% milk/TBST and detection was performed using an AP-conjugated Goat anti-Rabbit secondary antibody at a dilution of 1:10,000. NBT/BCIP was used as the substrate (Product # WB7105).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of NFkB p65 was performed by loading 40 µg of Jlat-full whole cell lysate in reducing sample buffer onto a 4-15% tris-glycine gel. Proteins were transferred to PVDF and then blocked in blocking buffer (TBST+5% non-fat milk) for one hour at room temperature. NFkB p65 was detected at approximately 65 kDa using a NFkB p65 Recombinant Rabbit Polyclonal Antibody (4-2HCLC) (Product # 710048) at a dilution of 1:1,000 overnight at 4°C on a rocking platform, followed by an Alexa Flour 488 conjugated Goat anti-Rabbit IgG Antibody (Product # A-11008) at a dilution of 1:3,000 for one hour at room temperature. Data courtesy of Antibody Data Exchange Program.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescent analysis of NF-kappa-B p65 was done on 70% confluent log phase TNF-Alpha treated HeLa cells (serum starved for 16 hours followed by treatment with 20 ng/mL TNF-Alpha for 1 hour). The cells were fixed with 4% paraformaldehyde for 15 minutes; permeabilized with 0.25% Triton X-100 for 10 minutes followed by blocking with 5% BSA for 1 hour at room temperature. The cells were incubated with NF-kappa-B p65 Recombinant Rabbit Polyclonal Antibody (Product # 710048) at 1 µg/mL in 1% BSA and incubated for 3 hours at room temperature and then labeled with Alexa Fluor 488 Goat anti-Rabbit IgG Secondary Antibody (Product # A-11008) at a dilution of 1:400 for 30 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 Alexa Fluor 594 Phalloidin (Product # A12381). Panel d shows nuclear localization of SMAD2 upon treatment; Panel e showing cytoplasmic localization in untreated HeLa cells. Panel f is a no primary antibody control. The images were captured at 20X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescence analysis of NFkB p65 was done on 70% confluent log phase ME-180 cells. The cells were either mock treated or treated with TNF-alpha (50 ng/mL for 20 min), fixed with 4% paraformaldehyde for 15 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were subsequently labeled with NFkB p65 (Green) Recombinant Rabbit Polyclonal Antibody (Product # 710048) at 1µg/mL in 0.1% BSA and incubated for 3 hours at room temperature 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. Nuclei (Blue) were stained with SlowFade® Gold Antifade Mountant DAPI (S36938). F-actin (Red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). TNFalpha induced nuclear translocation of NFkB, a downstream target in the TNFR1, TRADD and IKK alpha was observed in control cell line (panels a, e) and not in the TNFR1, TRADD and IKK alpha knockout (KO) cell lines (panels b-d; f-h). The images were captured at 40X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemistry analysis of NF-kappa-B (p65) showing staining in the cytoplasm and nucleus of paraffin-embedded human tonsil tissue (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature, washed with ddH2O and PBS, and then probed with a NF-kappa-B (p65) Recombinant Rabbit Polyclonal Antibody (Product # 710048) diluted in 3% BSA-PBS at a dilution of 1:20 overnight at 4°C in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using an HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemistry analysis of NF-kappa-B (p65) showing staining in the nucleus of paraffin-embedded mouse spleen tissue (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature, washed with ddH2O and PBS, and then probed with a NF-kappa-B (p65) Recombinant Rabbit Polyclonal Antibody (Product # 710048) diluted in 3% BSA-PBS at a dilution of 1:20 overnight at 4°C in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using an HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: ChIP- qPCR analysis of NF-kappa-B-p65 was performed with 3 µg/mL of the NF-kappa-B-p65 Recombinant Rabbit Polyclonal Antibody (Product # 710048) on sheared chromatin from 2 million HeLa cells treated with TNF-alpha (50 ng/mL for 1h) using the MAGnify Chromatin Immunoprecipitation System (Product # 49-2024). Normal rabbit IgG (3 µg/mL) was used as a negative IP control. The purified DNA from each ChIP sample was analyzed by StepOnePlus Real-Time PCR System (Product # 4376600) with primers for the promoter of active IL-8 and IL-6 gene, used as positive control targets, and the GAPDH gene, used as negative control target. Data is presented as fold enrichment of the antibody signal versus the negative control IgG using the comparative CT method.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Indirect ELISA analysis of NFkB p65 in HeLa cell lysate (1 µg/well) using a NFkB p65 Recombinant Rabbit Polyclonal Antibody (Product # 710048) and TMB (Product # SB01) as a substrate.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: 9 FIG Ghrelin inhibited AP-1 and NF-kappaB signaling pathway . ( A ) IF results revealed that ghrelin could inhibited the expression of AP-1 in PM-induced inflammation. ( B ) WB result showed that ghrelin significantly decreased the expression of P65 and incresed IkappaBalpha expression, indicating that classical NF-kappaB signaling pathway was inhibited (mean +- SEM; n = 3; Student's t -test; * P < 0.05, ** P < 0.01, *** P < 0.001 vs. control).

710048

Antibody data