Human/Mouse/Rat Phospho-JNK (T183/Y185) Antibody

Detection of Human and Mouse Phospho-JNK (T183/Y185) by Western Blot. Western blot shows lysates of HEK293 human embryonic kidney cell line and NIH-3T3 mouse embryonic fibroblast cell line untreated (-) or treated (+) with 100 J/m²UV-C for 30 minutes. PVDF membrane was probed with 0.5 µg/mL of Rabbit Anti-Human/Mouse/Rat Phospho-JNK (T183/Y185) Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1205), followed by HRP-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # HAF008). Specific bands were detected for Phospho-JNK (T183/Y185) at approximately 46 and 54 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 6.

JNK in Human Breast Cancer Tissue. JNK phosphorylated at T183/Y185 was detected in immersion fixed paraffin-embedded sections of human breast cancer tissue using Rabbit Anti-Human/Mouse/Rat Phospho-JNK (T183/Y185) Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1205) at 15 µg/mL overnight at 4 °C. Tissue was stained using the Anti-Rabbit HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS005) and counterstained with hematoxylin (blue). View our protocol for Chromogenic IHC Staining of immersion fixed paraffin-embedded Tissue Sections.

JNK in Human Prostate. JNK phosphorylated at T183/Y185 was detected in immersion fixed paraffin-embedded sections of human prostate array using Rabbit Anti-Human/Mouse/Rat Phospho-JNK (T183/Y185) Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1205) at 15 µg/mL overnight at 4 °C. Tissue was stained using the Anti-Rabbit HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS005) and counterstained with hematoxylin (blue). Lower panel shows a lack of labeling if primary antibodies are omitted and tissue is stained only with secondary antibody followed by incubation with detection reagents. View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.

Detection of Human Phospho-JNK (T183/Y185) by Simple Western^TM. Simple Western lane view shows lysates of HEK293T human embryonic kidney cell line untreated (-) or treated (+) with 100 J/m²ultraviolet light (UV) for 30 minutes, loaded at 0.2 mg/mL. Specific bands were detected for Phospho-JNK (T183/Y185) at approximately 56 and 46 kDa (as indicated) using 5 µg/mL of Rabbit Anti-Human/Mouse/Rat Phospho-JNK (T183/Y185) Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1205). This experiment was conducted under reducing conditions and using the 12-230 kDa separation system.*Non-specific interaction with the 230 kDa Simple Western standard may be seen with this antibody.

Detection of Mouse JNK1/2/3 by Western Blot Effect of p38MAPK (p38) inhibition on intracellular signaling following IR. Rats were pretreated with the carrier DMSO or BIRB796 (B-796) (5 mg/kg BW) for 1 hour and subjected to 1 hour of renal ischemia followed by different time points of reperfusion (15 min, 2 days, 7 days). Kidneys were harvested at given time points of reperfusion and total tissue lysates were used to determine activation pattern of MAPKs (p38MAPK, JNK, ERK) and the downstream target of p38MAPK (MK2) by phosphorylation specific antibodies. A representative immunoblot (A) and summary graphs (B, C) are shown. Results are given as mean ± SEM (n = 3). ** p < 0.01, ***p < 0.001 vs. sham-operated group; §p < 0.01, #p < 0.001 vs. IR-15 min group. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/24423080), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Mouse JNK1/2/3 by Western Blot Inhibition of JNK kinases prevents p66ShcS36 phosphorylation under oxidative stress.MEFs were treated with H2O2 (a) (1 mM, 30 min) or exposed to HR (hypoxia 90 min, reoxygenation 15 min) (b). Cell lysates were probed with the antibodies indicated to assess p66Shc phosphorylation at S36. Representative Western blots (individual experiment performed under the same experimental conditions and run on the same gel) and summary graphs from at least three independent experiments are shown. Signal intensity of control samples was arbitrarily set at 1. Mitochondrial ROS production was measured in p66Shc+/+ and p66Shc−/− MEFs after prooxidant treatment as described in Material and Methods (n ≥ 4) (c) and phospho gamma H2AX was detected by immunoblotting (d, lower panel) and summary graph is shown in above panel. SP: SP600125, JNK inhibitor, and BR: BR796, p38 inhibitor were applied 1 h prior to stress application. Original blot shown in (b) have been cropped and the full length blots are shown in the Supplementary Figure S1b Statistical analysis was performed using t-test or ANOVA (*p < 0.05 **p < 0.01, ***p < 0.001). Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/26868434), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Mouse JNK1/2/3 by Western Blot The impact of high glucose conditions on various down-stream signaling pathways.The status of various signaling pathways in retinal AC under various glucose conditions was determined as described in Methods. Levels of active Src (A), Akt (B), ERK (C), p38 (D), JNK1 (E), Fyn (F) and p65 (G) in retinal AC under different glucose conditions were determined by Western blot analysis from equal amounts of cell lysates. The total level for each protein is shown in the lower part of each panel. The quantitative assessment of the data for each blot is shown below the blot. Data are presented as mean ± SEM. n = 3; *p<0.05 (NG vs. HG and NG vs. NG+L-Glu). Levels of active Fyn (F) in retinal AC under different glucose conditions was determined by Western blot analysis of immunoprecipitates from equal amounts of retinal AC lysates. Image collected and cropped by CiteAb from the following open publication (https://dx.plos.org/10.1371/journal.pone.0103148), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Mouse JNK1/2/3 by Western Blot Effect of p38MAPK (p38) inhibition on intracellular signaling following IR. Rats were pretreated with the carrier DMSO or BIRB796 (B-796) (5 mg/kg BW) for 1 hour and subjected to 1 hour of renal ischemia followed by different time points of reperfusion (15 min, 2 days, 7 days). Kidneys were harvested at given time points of reperfusion and total tissue lysates were used to determine activation pattern of MAPKs (p38MAPK, JNK, ERK) and the downstream target of p38MAPK (MK2) by phosphorylation specific antibodies. A representative immunoblot (A) and summary graphs (B, C) are shown. Results are given as mean ± SEM (n = 3). ** p < 0.01, ***p < 0.001 vs. sham-operated group; §p < 0.01, #p < 0.001 vs. IR-15 min group. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/24423080), licensed under a CC-BY license. Not internally tested by R&D Systems.