ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free

Immunocytochemistry/ Immunofluorescence: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124]

ARNT-HIF-1-beta-Antibody-H1beta234-Immunocytochemistry-Immunofluorescence-NB100-124-img0011.jpg

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234)BSA Free [NB100-124]

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) [NB100-124] - Analysis of HIF-1 beta in HeLa nuclear extract using ARNT/HIF-1 beta antibody (H1beta234) [NB100-124]. Theoretical molecular weight 86.6 kDa. Observed molecular weight ~85 kDa.

Immunocytochemistry/ Immunofluorescence: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124]

Immunocytochemistry/Immunofluorescence: ARNT/HIF-1 beta Antibody (H1beta234) [NB100-124] - HeLa cells were fixed for 10 minutes using 10% formalin and then permeabilized for 5 minutes using 1X TBS + 0.5% Triton X-100. The cells were incubated with ARNT/HIF-1 beta antibody (H1beta234) [NB100-124] at 5 ug/mL overnight at 4C and detected with an anti-mouse DyLight 488 (Green) at a 1:500 dilution. Actin was detected with Phalloidin 568 (Red) at a 1:200 dilution. Nuclei were counterstained with DAPI (Blue). Cells were imaged using a 40X objective.

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] -

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] - PGE1 induces HIF-1 alpha protein accumulation in vascular-derived cells.Human aortic smooth muscle cells (HASMCs) (A & B), human umbilical vein endothelial cells (HUVECs) (C & D), & HEK293 cells (G) were exposed to 1 or 10 µM PGE1 under 20% O2 or 1% O2 conditions for 4 h. After treatment, cells were harvested & whole-cell lysates were subjected to immunoblot assay for HIF-1 alpha, HIF-1 beta, & beta-actin protein expression. Experiments were repeated thrice (A, C & G). Representative immunoblots are shown (A & C). Band intensities were analyzed densitometrically (B & D). Fold induction relative to lane 1 was plotted as mean ± S.D. ∗P < 0.05 compared with the control. HASMCs (E) & HUVECs (F) were exposed to 1 µM PGE1 for the indicated times under 20% O2 & were harvested for immunoblot assay for HIF-1 alpha protein. Experiments were repeated twice. Representative immunoblots are shown. (H) HASMCs & HUVECs were exposed to 10 µM PGE1 for 4 h under 20% O2 & were harvested for immunoblot assay for HIF-2 alpha protein. Experiments were repeated twice. Representative immunoblots are shown. I. HASMCs were exposed to 1 µM PGE1, lipo-PGE1 & PGE1-alfadex under 20% O2 conditions for 4 h. After treatment, cells were harvested & whole-cell lysates were subjected to immunoblot assay for HIF-1 alpha. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/24349900), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] -

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] - Effect of PGE1 on stability & synthesis of HIF-1 alpha.(A) Human aortic smooth muscle cells (HASMCs) & human umbilical vein endothelial cells (HUVECs) were exposed to 1 µM PGE1 or 100 µM DFX or incubated for 4 h, & CHX was added to a final concentration of 100 µM. The cells were incubated for 0 to 30 min, & whole-cell lysates were subjected to immunoblot assay using anti-HIF-1 alpha or -beta antibodies. (B) Serum-starved HASMCs were pretreated with no drug & 1 µM PGE1 for 30 min in Met-free medium. [35S]Met-Cys was added, & the cells were incubated for 60 min prior to preparation of cell lysates. Aliquots of 1 mg of the lysates were subjected to immunoprecipitation with anti-HIF-1 alpha antibody, separated by SDS-PAGE & exposed. Aliquots of 50 µg of the same lysate were separately subjected to immunoblotting analysis with anti-beta -actin antibody. (C) HASMCs & HUVECs were exposed to vehicle or 1 µM PGE1 for 4 h in the presence of 10 µM LY294002 (LY), 50 µM PD98059 (PD), or 5 µM GF109203X (GF). The cells were harvested & the whole-cell lysates were subjected to immunoblot assay for HIF-1 alpha & beta-actin protein expression. Experiments were repeated at least twice. Representative immunoblots are shown. (D) HASMCs were exposed to vehicle or 1 µM PGE1 for 12 h in the presence of 10 µM LY294002 (LY), 50 µM PD98059 (PD), or 5 µM GF109203X (GF). Cells were harvested & subjected to semi-quantitative RT-PCR for VEGF & GLUT1. Experiments were repeated three times. Fold induction relative to that under 20% O2 without PGE1 treatment was plotted. ∗P < 0.05 compared with the control (20%, PGE1 treatment without any kinase inhibitors). Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/24349900), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] -

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] - Effect of PGE1 on stability & synthesis of HIF-1 alpha.(A) Human aortic smooth muscle cells (HASMCs) & human umbilical vein endothelial cells (HUVECs) were exposed to 1 µM PGE1 or 100 µM DFX or incubated for 4 h, & CHX was added to a final concentration of 100 µM. The cells were incubated for 0 to 30 min, & whole-cell lysates were subjected to immunoblot assay using anti-HIF-1 alpha or -beta antibodies. (B) Serum-starved HASMCs were pretreated with no drug & 1 µM PGE1 for 30 min in Met-free medium. [35S]Met-Cys was added, & the cells were incubated for 60 min prior to preparation of cell lysates. Aliquots of 1 mg of the lysates were subjected to immunoprecipitation with anti-HIF-1 alpha antibody, separated by SDS-PAGE & exposed. Aliquots of 50 µg of the same lysate were separately subjected to immunoblotting analysis with anti-beta -actin antibody. (C) HASMCs & HUVECs were exposed to vehicle or 1 µM PGE1 for 4 h in the presence of 10 µM LY294002 (LY), 50 µM PD98059 (PD), or 5 µM GF109203X (GF). The cells were harvested & the whole-cell lysates were subjected to immunoblot assay for HIF-1 alpha & beta-actin protein expression. Experiments were repeated at least twice. Representative immunoblots are shown. (D) HASMCs were exposed to vehicle or 1 µM PGE1 for 12 h in the presence of 10 µM LY294002 (LY), 50 µM PD98059 (PD), or 5 µM GF109203X (GF). Cells were harvested & subjected to semi-quantitative RT-PCR for VEGF & GLUT1. Experiments were repeated three times. Fold induction relative to that under 20% O2 without PGE1 treatment was plotted. ∗P < 0.05 compared with the control (20%, PGE1 treatment without any kinase inhibitors). Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/24349900), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] -

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] - PGE1 induces HIF-1 alpha protein accumulation in vascular-derived cells.Human aortic smooth muscle cells (HASMCs) (A & B), human umbilical vein endothelial cells (HUVECs) (C & D), & HEK293 cells (G) were exposed to 1 or 10 µM PGE1 under 20% O2 or 1% O2 conditions for 4 h. After treatment, cells were harvested & whole-cell lysates were subjected to immunoblot assay for HIF-1 alpha, HIF-1 beta, & beta-actin protein expression. Experiments were repeated thrice (A, C & G). Representative immunoblots are shown (A & C). Band intensities were analyzed densitometrically (B & D). Fold induction relative to lane 1 was plotted as mean ± S.D. ∗P < 0.05 compared with the control. HASMCs (E) & HUVECs (F) were exposed to 1 µM PGE1 for the indicated times under 20% O2 & were harvested for immunoblot assay for HIF-1 alpha protein. Experiments were repeated twice. Representative immunoblots are shown. (H) HASMCs & HUVECs were exposed to 10 µM PGE1 for 4 h under 20% O2 & were harvested for immunoblot assay for HIF-2 alpha protein. Experiments were repeated twice. Representative immunoblots are shown. I. HASMCs were exposed to 1 µM PGE1, lipo-PGE1 & PGE1-alfadex under 20% O2 conditions for 4 h. After treatment, cells were harvested & whole-cell lysates were subjected to immunoblot assay for HIF-1 alpha. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/24349900), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] -

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] - Differential involvement of EP receptors in PGE1-induced HIF-1 alpha protein accumulation.(A) Expression of EP1, EP2, EP3, & EP4 receptors in human aortic smooth muscle cells (HASMCs), human umbilical vein endothelial cells (HUVECs), & cells of the neuroblastoma cell line SH-SY5Y. HASMCs, HUVECs, & SH-SY5Y cells were cultured under 20% O2 & harvested for semi-quantitative RT-PCR for EP1-4 receptors. Experiments were repeated at least three times in triplicate. Fold induction relative to expression in SH-SY5Y cells was plotted as mean ± S.D. HASMCs & HUVECs were exposed to 1 µM of EP-receptor-specific agonists (ONO-DI-004 for EP1, ONO-AE1-259-01 for EP2, ONO-AE-248 for EP3, & ONO-AE1-329 for EP4) for 4 h (B). HASMCs & HUVECs were exposed to 1 µM PGE1 with or without 1 µM EP-receptor-specific antagonists (ONO-8713 against EP1, ONO-AE3-240 against EP3, & ONO-AE2-227 against EP4) for 4 h (C). The cells were harvested & the whole-cell lysates were subjected to immunoblot assay for HIF-1 alpha & beta-actin protein expression. Experiments were repeated twice. Representative immunoblots are shown. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/24349900), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] -

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] - HIF was not the only factor stabilizing activated EGFR in VHL-deficient ccRCC cells.A. Western blot analysis of 786-VHL & 786-mock cells stably expressing shRNA constructs. For HIF2 alpha & HIF1 beta analysis, nuclear extracts were generated & analyzed. Anti-HA blot detected HA-VHL. B. 786-VHL cells expressing SCR (control), HIF2a-566 & HIF2a-1631 (shRNA constructs against HIF2 alpha) were treated with EGF & analyzed as described in Fig. 1A. C. 786-mock cells expressing SCR (control sequence), HIF2a-566 & HIF2a-1631 (shRNA constructs against HIF2 alpha) were treated with EGF & analyzed as described in Fig. 2B. D. The EGFR signals in Fig. 2B & 2C were normalized over Vinculin via densitometry & plotted over time. Means & SDs of three separate experiments were shown. Image collected & cropped by CiteAb from the following publication (https://dx.plos.org/10.1371/journal.pone.0023936), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Immunocytochemistry/ Immunofluorescence: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] -

Immunocytochemistry/ Immunofluorescence: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] - Co-localization of HIF-2 alpha with HIF-1 beta & RNAPII. (a) Immunofluorescence detecting indicated proteins using antibodies labelled with Alexa Fluor 555 (red, pseudocolour) & Alexa Fluor 488 (green, pseudocolour). ‘Merge’ is the red image superimposed onto the green image of the co-stained nuclear proteins. ‘Coloc’ is the co-localization channel calculated using ImageJ plugin Co-localization Threshold. White indicates pixels where both red & green signal is found (i.e. co-localization). ‘Overlay’ is the co-localization image superimposed onto the merged image. Inlay is the magnified region (white square). White arrows highlight regions of co-localization. Scale bar, 5 μm. Abbreviations: RNAPII, RNA Polymerase II phospho-serine 5. (b) Immunofluorescence images were analysed using ImageJ plugin Co-localization Threshold with use of the Costes et al. [31] method to automatically create a threshold prior to calculating the Mander's coefficient for both proteins. The results are given as, for example, the percentage of protein A (HIF-2 alpha) that co-localized with protein B (HIF-1 beta or RNAPII) & vice versa. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/27655733), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] -

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] - Differential involvement of EP receptors in PGE1-induced HIF-1 alpha protein accumulation.(A) Expression of EP1, EP2, EP3, & EP4 receptors in human aortic smooth muscle cells (HASMCs), human umbilical vein endothelial cells (HUVECs), & cells of the neuroblastoma cell line SH-SY5Y. HASMCs, HUVECs, & SH-SY5Y cells were cultured under 20% O2 & harvested for semi-quantitative RT-PCR for EP1-4 receptors. Experiments were repeated at least three times in triplicate. Fold induction relative to expression in SH-SY5Y cells was plotted as mean ± S.D. HASMCs & HUVECs were exposed to 1 µM of EP-receptor-specific agonists (ONO-DI-004 for EP1, ONO-AE1-259-01 for EP2, ONO-AE-248 for EP3, & ONO-AE1-329 for EP4) for 4 h (B). HASMCs & HUVECs were exposed to 1 µM PGE1 with or without 1 µM EP-receptor-specific antagonists (ONO-8713 against EP1, ONO-AE3-240 against EP3, & ONO-AE2-227 against EP4) for 4 h (C). The cells were harvested & the whole-cell lysates were subjected to immunoblot assay for HIF-1 alpha & beta-actin protein expression. Experiments were repeated twice. Representative immunoblots are shown. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/24349900), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Immunocytochemistry/ Immunofluorescence: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] -

Immunocytochemistry/ Immunofluorescence: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] - Sub-nuclear localization of HIF-1 alpha & HIF-2 alpha. (a) HeLa cells ectopically expressing HIF-1 alpha & HIF-2 alpha EGFP fusions compared with endogenous HIF-1 alpha & HIF-2 alpha labelled using immunostaining. Images of HIF-1 alpha were taken following DMOG treatment (6 h; 0.5 mM). Scale bar, 5 μm. (b) HeLa cells transiently transfected with plasmids encoding (i) clover-HIF-2 alpha (green, pseudocolour), (ii) dsRED-HIF-2 alpha (red, pseudocolour), (iii) HIF-2 alpha-venus (yellow pseudocolour) & (iv) Halotag-HIF-2 alpha (green, pseudocolour). The cells expressing Halotag-HIF-2 alpha were labelled with the fluorescent Oregon Green Halotag ligand (HL-OregonGreen; Promega, WI, USA) to visualize the fusion protein. (c) Confocal images of C2C12 (mouse myoblast; top) & HEK293T (Human embryonic kidney cells; bottom) cells ectopically expressing EGFP-HIF-2 alpha. Scale bar, 5 μm. (d) HeLa cells transiently transfected with EGFP-HIF-2 alpha were imaged with a CCD camera. One thousand frames were acquired per cell in normoxia, hypoxia (1% v/v O2, 16 h) or following treatment with DMOG (0.5 mM, 6 h). The average (±s.d.) number of speckles per nucleus in each condition was 64 ± 49 (n = 25), 44 ± 24 (n = 24) & 96 ± 33 (n = 22), respectively. Mean of the sample data represented by the red dashed line. (e) Using the images from (d) the average speckle area per nucleus over the 1000 frames. The mean values (±s.d.) for each condition were 0.24 ± 0.09 µm (n = 25), 0.21 ± 0.07 µm (n = 24) & 0.27 ± 0.09 µm (n = 22), respectively. The mean values for hypoxia & DMOG were compared with the normoxic values (independent t-test, significance value set at 5%). Mean of the sample data represented by the red dashed line. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/27655733), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] -

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] - PGE1 induces HIF-1 alpha protein accumulation in vascular-derived cells.Human aortic smooth muscle cells (HASMCs) (A & B), human umbilical vein endothelial cells (HUVECs) (C & D), & HEK293 cells (G) were exposed to 1 or 10 µM PGE1 under 20% O2 or 1% O2 conditions for 4 h. After treatment, cells were harvested & whole-cell lysates were subjected to immunoblot assay for HIF-1 alpha, HIF-1 beta, & beta-actin protein expression. Experiments were repeated thrice (A, C & G). Representative immunoblots are shown (A & C). Band intensities were analyzed densitometrically (B & D). Fold induction relative to lane 1 was plotted as mean ± S.D. ∗P < 0.05 compared with the control. HASMCs (E) & HUVECs (F) were exposed to 1 µM PGE1 for the indicated times under 20% O2 & were harvested for immunoblot assay for HIF-1 alpha protein. Experiments were repeated twice. Representative immunoblots are shown. (H) HASMCs & HUVECs were exposed to 10 µM PGE1 for 4 h under 20% O2 & were harvested for immunoblot assay for HIF-2 alpha protein. Experiments were repeated twice. Representative immunoblots are shown. I. HASMCs were exposed to 1 µM PGE1, lipo-PGE1 & PGE1-alfadex under 20% O2 conditions for 4 h. After treatment, cells were harvested & whole-cell lysates were subjected to immunoblot assay for HIF-1 alpha. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/24349900), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] -

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] - PGE1 induces HIF-1 alpha protein accumulation in vascular-derived cells.Human aortic smooth muscle cells (HASMCs) (A & B), human umbilical vein endothelial cells (HUVECs) (C & D), & HEK293 cells (G) were exposed to 1 or 10 µM PGE1 under 20% O2 or 1% O2 conditions for 4 h. After treatment, cells were harvested & whole-cell lysates were subjected to immunoblot assay for HIF-1 alpha, HIF-1 beta, & beta-actin protein expression. Experiments were repeated thrice (A, C & G). Representative immunoblots are shown (A & C). Band intensities were analyzed densitometrically (B & D). Fold induction relative to lane 1 was plotted as mean ± S.D. ∗P < 0.05 compared with the control. HASMCs (E) & HUVECs (F) were exposed to 1 µM PGE1 for the indicated times under 20% O2 & were harvested for immunoblot assay for HIF-1 alpha protein. Experiments were repeated twice. Representative immunoblots are shown. (H) HASMCs & HUVECs were exposed to 10 µM PGE1 for 4 h under 20% O2 & were harvested for immunoblot assay for HIF-2 alpha protein. Experiments were repeated twice. Representative immunoblots are shown. I. HASMCs were exposed to 1 µM PGE1, lipo-PGE1 & PGE1-alfadex under 20% O2 conditions for 4 h. After treatment, cells were harvested & whole-cell lysates were subjected to immunoblot assay for HIF-1 alpha. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/24349900), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] -

Western Blot: ARNT/HIF-1 beta Antibody (H1beta234) - BSA Free [NB100-124] - PGE1 induces HIF-1 alpha protein accumulation in vascular-derived cells.Human aortic smooth muscle cells (HASMCs) (A & B), human umbilical vein endothelial cells (HUVECs) (C & D), & HEK293 cells (G) were exposed to 1 or 10 µM PGE1 under 20% O2 or 1% O2 conditions for 4 h. After treatment, cells were harvested & whole-cell lysates were subjected to immunoblot assay for HIF-1 alpha, HIF-1 beta, & beta-actin protein expression. Experiments were repeated thrice (A, C & G). Representative immunoblots are shown (A & C). Band intensities were analyzed densitometrically (B & D). Fold induction relative to lane 1 was plotted as mean ± S.D. ∗P < 0.05 compared with the control. HASMCs (E) & HUVECs (F) were exposed to 1 µM PGE1 for the indicated times under 20% O2 & were harvested for immunoblot assay for HIF-1 alpha protein. Experiments were repeated twice. Representative immunoblots are shown. (H) HASMCs & HUVECs were exposed to 10 µM PGE1 for 4 h under 20% O2 & were harvested for immunoblot assay for HIF-2 alpha protein. Experiments were repeated twice. Representative immunoblots are shown. I. HASMCs were exposed to 1 µM PGE1, lipo-PGE1 & PGE1-alfadex under 20% O2 conditions for 4 h. After treatment, cells were harvested & whole-cell lysates were subjected to immunoblot assay for HIF-1 alpha. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/24349900), licensed under a CC-BY license. Not internally tested by Novus Biologicals.