Nlk Antibody (B-5) is a high quality monoclonal Nlk antibody (also designated Nlk antibody) suitable for the detection of the Nlk protein of mouse, rat and human origin. Nlk Antibody (B-5) is available as both the non-conjugated anti-Nlk antibody form, as well as multiple conjugated forms of anti-Nlk antibody, including agarose, HRP, PE, FITC and multiple Alexa Fluor® conjugates. The activation of signal transduction pathways by growth factors, hormones and neurotransmitters is mediated through two closely related MAP kinases, p44 and p42, designated extracellular-signal related kinase 1 (ERK 1) and ERK 2 (1-4), respectively. ERK proteins are regulated by dual phosphorylation at specific tyrosine and threonine sites mapping within a characteristic Thr-Glu-Tyr motif. Phosphorylation at both Thr 183 and Tyr 185 is required for full enzymatic activation. In response to activation, MAP kinases phosphorylate downstream components on serine and threonine. Nlk, or nemo-like kinase, is a murine homolog of the Drosophila nemo (nmo) gene. Nlk and Nmo have sequence homology to both the ERK MAP kinases and the cyclin dependent kinases. Nlk is a nuclear protein with the ability to autophosphorylate.
Background References
1. Li SZ. et. al. Phosphorylation of MAVS/VISA by Nemo-like kinase (NLK) for degradation regulates the antiviral innate immune response. Nat Commun. 2019 Jul
2. Fleskens V. et. al. Nemo-like Kinase Drives Foxp3 Stability and Is Critical for Maintenance of Immune Tolerance by Regulatory T Cells. Cell Rep. 2019 Mar
Western blot analysis of NLK on different lysates with Rabbit anti-NLK antibody (HA500033) at 1/2,000 dilution.
Lane 1: Jurkat (Human T-lymphoblastic cells) cell lysate Lane 2: Caco-2 (Human colorectal adenocarcinoma cell) cell lysate
Lysates/proteins at 10 µg/Lane. Exposure time: 46 seconds; ECL: K1801
Blocking: 5% NFDM/TBST, 1 hour at room temperature Primary antibody: HA500033, 1/2,000 in primary antibody dilution buffer (K1803), overnight at 4 ℃ Secondary antibody: Goat anti-Rabbit IgG-HRP (HA1001), 1/50,000 in 5% NFDM/TBST, 1 hour at room temperature
Predicted band size: 58 kDa Observed band size: 58 kDa
ICC staining of NLK in MG-63 cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with the primary antibody (HA500033, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).
ICC staining of NLK in SW620 cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with the primary antibody (HA500033, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).
Immunohistochemical analysis of paraffin-embedded human uterus tissue using anti-NLK antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA500033, 1/200) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
Immunohistochemical analysis of paraffin-embedded human small intestine tissue using anti-NLK antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA500033, 1/200) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
Immunohistochemical analysis of paraffin-embedded human breast carcinoma tissue using anti-NLK antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA500033, 1/200) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
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