Chromatin Immunoprecipitation, Chromatin Immunoprecipitation (ChIP), CyTOF-ready, ELISA, Flow Cytometry, Gel Super Shift Assays, Immunoassay, Immunoblotting, Immunocytochemistry/ Immunofluorescence, Immunohistochemistry, Immunohistochemistry-Frozen, Immunohistochemistry-Paraffin, Immunoprecipitation, In vitro assay, Knockdown Validated, Knockout Validated, Ligand Activation, Proximity Ligation Assay, Tissue Culture Substratum, Western Blot

Label

Unconjugated

Antibody Source

Monoclonal Mouse IgG_2B Clone # H1alpha67

Format

Azide and BSA Free

Concentration

1 mg/ml

View all HIF-1 alpha/HIF1A Antibodies »

Product Specifications

Immunogen

This HIF-1 alpha Antibody (H1alpha67) was developed against a fusion protein containing amino acids 432 - 528 of human HIF-1 alpha [Uniprot# Q16665].

Reactivity Notes

Xenopus reactivity was reported in scientific literature (PMID: 18303027). Please note that this antibody is reactive to Mouse and derived from the same host, Mouse. Additional Mouse on Mouse blocking steps may be required for IHC and ICC experiments. Please contact Technical Support for more information. Canine reactivity reported in scientific literature (PMID: 24153191, 29974500). Feline reactivity reported in scientific literature (PMID: 30419801).

Localization

HIF-1 is a nuclear protein

Clonality

Monoclonal

Host

Mouse

Isotype

IgG_2B

Theoretical MW

93 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Scientific Data Images for HIF-1 alpha Antibody (H1alpha67) - Azide and BSA Free

Detection of HIF-1 alpha by Western Blot in Multiple Cells Lines with Various Treatments

Upregulation of HIF-1alpha in human PCa. Protein expression of HIF-1alpha, VEGF, and GLUT4 were examined with western blot, in PC-3, DU145, and LNCaP cells after various treatments as indicated. Data are expressed as mean +/- SD of seven independent experiments. $p < 0.05 versus RWPE-1 or BPH1 cells or normal tissue. *p < 0.05 versus control group. #p < 0.05 versus si-HIF-1alpha or DDP group. Original blots are shown in Supplementary Figure 5. C: Ctrl; D: DDP; S: si-HIF-1alpha; D/S: DDP/si-HIF-1alpha. Image collected and cropped by CiteAb from the following publication (https://www.nature.com/articles/s41598-017-07973-4), licensed under a CC-BY license. Image from the standard format of this antibody.

Staining of HIF-1 alpha in DFOA Treated and non-Treated HeLa Cells

HIF-1 alpha was detected in immersion fixed HeLa human cervical epithelial carcinoma cell line treated with DFOA using 1 ug/mL of mouse anti-HIF-1 alpha monoclonal antibody NB100-105. Cells were stained using a donkey anti-rabbit secondary antibody and counterstained with DAPI (blue). Image from the standard format of this antibody.

Immunohistological Staining of HIF-1 alpha in Paraffin Embedded Human Glioblastoma Multiforme

Analysis using the biotin conjugate of NB100-105. Staining of human glioblastoma multiforme.

Flow Cytometry of Myeloma Cells Stained with Alexa Fluor 488 Conjugated HIF-1 alpha Antibody

Analysis using the Alexa Fluor (R) 488 conjugate of NB100-105. Staining of HIF-1 alpha in multiple myeloma cells: H929 cells (0.5x10^6) were stained with Alexa Fluor 488 (R) conjugated HIF-1 alpha antibody (NB100-105AF488). Image courtesy of Dr. Barbara Muz at Washington University in St. Louis School of Medicine.

Western Blot Analysis of HIF-1 alpha in Human Hepatocytes

Analysis of HIF-1 alpha in human hepatocytes from cancer patient using anti-HIF-1 alpha antibody NB100-105. Image from verified customer review. Image from the standard format of this antibody.

Use of HRP Conjugated HIF-1 alpha Antibody in Western Blotting of COS-7 Nuclear Extracts

Analysis using the HRP conjugate of NB100-105. Detection of 50ug cobalt chloride induced COS-7 nuclear extracts (NB800-PC26) using NB100-105.

Western Blot Analysis of HIF-1 alpha in Caki-1 Cell Lysates

HIF-1 alpha induction by CoCl2 on Caki-1 cell lysate. Image from verified customer review. Image from the standard format of this antibody.

Western Blot Analysis of HIF-1 alpha in Nuclear-Enriched and Whole Cell Extracts from Non-Diabetic and T2D Donor Islets

HIF1alpha/PFKFB3 is upregulated in beta-cells of HIP rats and humans with type 2 diabetes. Representative Western blot of PFKFB3 and HIF1alpha levels in nuclear-enriched- and whole cell extracts from non-diabetic (ND) and T2D donor islets. Data are presented as mean +/- SEM, n = 3 independent biological samples for each group. Statistical significance was analyzed by Student t-test (*p < 0.05, ***p < 0.001) Image collected and cropped by CiteAb from the following publication (https://www.nature.com/articles/s41467-019-10444-1) licensed under a CC-BY license. Image from the standard format of this antibody.

Western Blot of HIF-1 alpha in CD11c+ Cells from C57BL/6 Mice and Densitometric Analysis

L. donovani infection induces HIF-1 alpha expression in CD11chi splenic DCs in an IRF-5 dependent manner. Mice were infected with 2x107 amastigotes intravenously. Immunoblot analysis of HIF-1 alpha expression in CD11c+ cells from C57BL/6 mice (upper panel) and densitometric analysis normalized to beta-actin expression and expressed as fold increase to results obtained with naive mice (lower panel). Image collected and cropped by CiteAb from the following publication (https://dx.plos.org/10.1371/journal.ppat.1004938), licensed under a CC-BY license. Image from the standard format of this antibody.

Detection of HIF-1 alpha in WT and HIF-1 alpha Knockout Mouse Cells by Western Blot

Naive CD4 T cells from WT, VHL-deficient (Vhl KO), or HIF-1 alpha-deficient (HIF-1 alpha KO) mice were differentiated under IL-22-skewing conditions for a total of 60 h. Some cells remained at normoxia for the duration of the culture (N); others were at normoxia for 35 h and then hypoxia (1% O2) for 24 h (H). At 60 h, nuclear extracts were harvested, and HIF-1 alpha and Lamin B1 levels were analyzed by Western blot. Image from verified customer review. Image from the standard format of this antibody.

Western Blot Detection of HIF-1 alpha in Nonmalignant and PCa Cell Lines

Upregulation of HIF-1alpha in human Pca. HIF-1alpha protein was detected by western blot in nonmalignant (RWPE-1 and BPH1) and PCa cell lines (PC-3, DU145, LNCaP, and 22RV1) as indicated. Image collected and cropped by CiteAb from the following publication (https://www.nature.com/articles/s41598-017-07973-4), licensed under a CC-BY license. Image from the standard format of this antibody.

Immunohistological Detection of HIF-1 alpha in PDAC and Adjacent Normal Pancreatic Tissue

HIF-1alpha expression in PDAC and adjacent normal pancreatic tissue. PDAC with weak (intensity 1) nuclear and cytoplasmic HIF-1alpha staining (left). PDAC with strong (intensity 3) nuclear and moderate (intensity 2) cytoplasmic HIF-1alpha staining (right)

Immunohistochemical Staining of HIF-1 alpha in Human Kidney

Staining of HIF-1 alpha in human kidney using NB100-105. Renal tubular epithelium showed moderate membranous, cytoplasmic and nuclear staining, and glomeruli showed faint to moderate nuclear staining. Image from the standard format of this antibody.

Flow Cytometry of H929 Cells Stained with Alexa Fluor 488 Conjugated HIF-1 alpha Antibody

Analysis using the Alexa Fluor (R) 488 conjugate of NB100-105. Staining of HIF-1 alpha in H929 cells using HIF-1 alpha antibody. Image from verified customer review.

Knockout Validation of HIF-1 alpha Antibody in HeLa and HIF-1 Knockout HeLa Cells

HIF-1 alpha was detected in immersion fixed DFO treated Hela cells (left) but was not detected in HIF-1 knockout HeLa cells (right) using Mouse Anti-human HIF-1 alpha monoclonal antibody (Catalog #NB100-105) at 25 ug/mL for 3 hours at room temperature. Cells were stained using a NorthernLights (TM) 557-conjugated Donkey Anti-Mouse IgG Secondary Antibody (red; Catalog # NL007) and counterstained with DAPI (blue). Specific staining was localized to nuclei. Image from the standard format of this antibody.

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Applications for HIF-1 alpha Antibody (H1alpha67) - Azide and BSA Free

Application

Recommended Usage

Chromatin Immunoprecipitation

1 - 5 ug/IP. Use reported in scientific literature

Chromatin Immunoprecipitation (ChIP)

1-5 ug/IP

ELISA

1:100 - 1:2000. Use reported in scientific literature (PMID 20042684)

Flow Cytometry

1:10 - 1:1000

Gel Super Shift Assays

1:1 - 1:100. Use reported in scientific literature (PMID 22411794)

Immunoassay

reported in scientific literature (PMID 26147748)

Immunoblotting

reported in multiple pieces of scientific literature

Immunocytochemistry/ Immunofluorescence

1:50

Immunohistochemistry-Frozen

1:20 - 1:50

Immunohistochemistry-Paraffin

1:20 - 1:50

Immunoprecipitation

1:10 - 1:500

In vitro assay

reported in multiple pieces of scientific literature

Knockout Validated

reported in scientific literature (PMID 26861754)

Ligand Activation

reported in scientific literature (PMID 26147748)

Proximity Ligation Assay

reported in scientific literature (PMID 27595394)

Application Notes

In WB, a band can be seen at 120 kDa representing HIF-1 alpha in induced tissues and cells. Multiple bands may be seen at 100-120 kDa representing post-translational modification of HIF-1 alpha. For WB, testing on nuclear extracts is recommended. We recommend the use of a highly sensitive ECL reagent, such as West Pico PLUS, for Western blot detection.

Formulation, Preparation, and Storage

Purification

Protein G purified

Formulation

PBS

Format

Azide and BSA Free

Preservative

No Preservative

Concentration

1 mg/ml

Shipping

The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles.

Background: HIF-1 alpha/HIF1A

Hypoxia contributes to the pathophysiology of human disease, including myocardial and cerebral ischemia, cancer, pulmonary hypertension, congenital heart disease and chronic obstructive pulmonary disease (1). In cancer and particularly solid tumors, hypoxia plays a critical role in the regulation of genes involved in stem cell renewal, epithelial to mesenchymal transition (EMT), metastasis and angiogenesis. In the tumor microenvironment (TME), hypoxia influences the properties and function of stromal cells (e.g., fibroblasts, endothelial and immune cells) and is a strong determinant of tumor progression (2,3).

HIF-1 or hypoxia inducible factor 1 (predicted molecular weight 93kDa), is a transcription factor commonly referred to as a "master regulator of the hypoxic response" for its central role in the regulation of cellular adaptations to hypoxia. In its active form under hypoxic conditions, HIF-1 is stabilized by the formation of a heterodimer of HIF-1 alpha and ARNT/HIF-1 beta subunits. Nuclear HIF-1 engages p300/CBP for binding to hypoxic response elements (HREs). This process induces transcription and regulation of genes including EPO, VEGF, iNOS2, ANGPT1 and OCT4 (4,5).

Under normoxic conditions, the HIF-1 alpha subunit is rapidly targeted and degraded by the ubiquitin proteasome system. This process is mediated by prolyl hydroxylase domain enzymes (PHDs), which catalyze the hydroxylation of key proline residues (Pro-402 and Pro-564) within the oxygen-dependent degradation domain of HIF-1 alpha. Once hydroxylated, HIF-1 alpha binds the von Hippel-Lindau tumor suppressor protein (pVHL) for subsequent ubiquitination and proteasomal degradation (4). pVHL dependent regulation of HIF-1 alpha plays a role in normal physiology and disease states. Regulation of HIF-1 alpha by pVHL is critical for the suppressive function of FoxP3+ regulatory Tcells (6). Repression of pVHL expression in chronic lymphocytic leukemia (CLL) B cells leads to HIF-1 alpha stabilization and increased VEGF secretion (7).

References

1. Semenza, G. L., Agani, F., Feldser, D., Iyer, N., Kotch, L., Laughner, E., & Yu, A. (2000). Hypoxia, HIF-1, and the pathophysiology of common human diseases. Advances in Experimental Medicine and Biology.

2. Muz, B., de la Puente, P., Azab, F., & Azab, A. K. (2015). The role of hypoxia in cancer progression, angiogenesis, metastasis, and resistance to therapy. Hypoxia. https://doi.org/10.2147/hp.s93413

3. Huang, Y., Lin, D., & Taniguchi, C. M. (2017). Hypoxia inducible factor (HIF) in the tumor microenvironment: friend or foe? Science China Life Sciences. https://doi.org/10.1007/s11427-017-9178-y

4. Koyasu, S., Kobayashi, M., Goto, Y., Hiraoka, M., & Harada, H. (2018). Regulatory mechanisms of hypoxia-inducible factor 1 activity: Two decades of knowledge. Cancer Science. https://doi.org/10.1111/cas.13483

5. Dengler, V. L., Galbraith, M. D., & Espinosa, J. M. (2014). Transcriptional regulation by hypoxia inducible factors. Critical Reviews in Biochemistry and Molecular Biology. https://doi.org/10.3109/10409238.2013.838205

6. Lee, J. H., Elly, C., Park, Y., & Liu, Y. C. (2015). E3Ubiquitin Ligase VHL Regulates Hypoxia-Inducible Factor-1 alpha to Maintain Regulatory T Cell Stability and Suppressive Capacity. Immunity. https://doi.org/10.1016/j.immuni.2015.05.016

7. Ghosh, A. K., Shanafelt, T. D., Cimmino, A., Taccioli, C., Volinia, S., Liu, C. G., ... Kay, N. E. (2009). Aberrant regulation of pVHL levels by microRNA promotes the HIF/VEGF axis in CLL B cells. Blood. https://doi.org/10.1182/blood-2008-10-185686

Long Name

Hypoxia Inducible Factor 1 Subunit Alpha

Alternate Names

BHLHE78, HIF 1A, HIF-1a, HIF1 alpha, HIF1A, MOP1, PASD8

Gene Symbol

HIF1A

Additional HIF-1 alpha/HIF1A Products

Product Documents for HIF-1 alpha Antibody (H1alpha67) - Azide and BSA Free

Product Datasheets

Download Product Datasheets

COA

SDS

Download SDS

Product Specific Notices for HIF-1 alpha Antibody (H1alpha67) - Azide and BSA Free

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

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HIF-1 alpha Antibody (H1alpha67) - Azide and BSA Free

Novus Biologicals, part of Bio-Techne | Catalog # NBP2-80760

Key Product Details

Validated by

Species Reactivity

Applications

Label

Antibody Source

Format

Concentration

View all HIF-1 alpha/HIF1A Antibodies »

Product Specifications

Immunogen

Reactivity Notes

Localization

Clonality

Host

Isotype

Theoretical MW

Scientific Data Images for HIF-1 alpha Antibody (H1alpha67) - Azide and BSA Free

Detection of HIF-1 alpha by Western Blot in Multiple Cells Lines with Various Treatments

Staining of HIF-1 alpha in DFOA Treated and non-Treated HeLa Cells

Immunohistological Staining of HIF-1 alpha in Paraffin Embedded Human Glioblastoma Multiforme

Flow Cytometry of Myeloma Cells Stained with Alexa Fluor 488 Conjugated HIF-1 alpha Antibody

Western Blot Analysis of HIF-1 alpha in Human Hepatocytes

Use of HRP Conjugated HIF-1 alpha Antibody in Western Blotting of COS-7 Nuclear Extracts

Western Blot Analysis of HIF-1 alpha in Caki-1 Cell Lysates

Western Blot Analysis of HIF-1 alpha in Nuclear-Enriched and Whole Cell Extracts from Non-Diabetic and T2D Donor Islets

Western Blot of HIF-1 alpha in CD11c+ Cells from C57BL/6 Mice and Densitometric Analysis

Detection of HIF-1 alpha in WT and HIF-1 alpha Knockout Mouse Cells by Western Blot

Western Blot Detection of HIF-1 alpha in Nonmalignant and PCa Cell Lines

Immunohistological Detection of HIF-1 alpha in PDAC and Adjacent Normal Pancreatic Tissue

Immunohistochemical Staining of HIF-1 alpha in Human Kidney

Flow Cytometry of H929 Cells Stained with Alexa Fluor 488 Conjugated HIF-1 alpha Antibody

Knockout Validation of HIF-1 alpha Antibody in HeLa and HIF-1 Knockout HeLa Cells

Applications for HIF-1 alpha Antibody (H1alpha67) - Azide and BSA Free

Chromatin Immunoprecipitation

Chromatin Immunoprecipitation (ChIP)

ELISA

Flow Cytometry

Gel Super Shift Assays

Immunoassay

Immunoblotting

Immunocytochemistry/ Immunofluorescence

Immunohistochemistry-Frozen

Immunohistochemistry-Paraffin

Immunoprecipitation

In vitro assay

Knockout Validated

Ligand Activation

Proximity Ligation Assay

Formulation, Preparation, and Storage

Purification

Formulation

Format

Preservative

Concentration

Shipping

Stability & Storage

Background: HIF-1 alpha/HIF1A

Long Name

Alternate Names

Gene Symbol

Additional HIF-1 alpha/HIF1A Products

Product Documents for HIF-1 alpha Antibody (H1alpha67) - Azide and BSA Free

Product Datasheets

COA

Certificate of Analysis

SDS

Product Specific Notices for HIF-1 alpha Antibody (H1alpha67) - Azide and BSA Free