Key Product Details

Validated by

Biological Validation

Species Reactivity

Validated:

Human

Cited:

Human, Mouse, Rat

Applications

Validated:

Immunohistochemistry, Western Blot, Intracellular Staining by Flow Cytometry, Immunocytochemistry, Chromatin Immunoprecipitation (ChIP), CyTOF-ready

Cited:

Immunohistochemistry, Immunohistochemistry-Paraffin, Western Blot, ELISA, Neutralization, Immunocytochemistry, Immunoprecipitation, ELISA Development, ELISA Standard, In vivo assay, Immunodepletion

Label

Unconjugated

Antibody Source

Monoclonal Mouse IgG2B Clone # 115603
View all HMGB1/HMG-1 Primary Antibodies »

Product Specifications

Immunogen

E. coli-derived recombinant human HMGB1/HMG-1
Gly2-Glu215 (predicted)
Accession # P09429

Specificity

Detects human HMGB1/HMG-1 in direct ELISAs and Western blots.

Clonality

Monoclonal

Host

Mouse

Isotype

IgG2B

Scientific Data Images for Human HMGB1/HMG‑1 Antibody

Detection of Human HMGB1/HMG-1 antibody by Western Blot.

Detection of Human HMGB1/HMG‑1 by Western Blot.

Western blot shows lysates of HepG2 human hepatocellular carcinoma cell line, Jurkat human acute T cell leukemia cell line, and HEK293 human embryonic kidney cell line. PVDF membrane was probed with 0.05 µg/mL of Mouse Anti-Human HMGB1/HMG-1 Monoclonal Antibody (Catalog # MAB1690) followed by HRP-conjugated Anti-Mouse IgG Secondary Antibody (Catalog # HAF018). A specific band was detected for HMGB1/HMG-1 at approximately 26 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.
Detection of HMGB1/HMG-1-regulated Genes antibody by Chromatin Immunoprecipitation.

Detection of HMGB1/HMG‑1-regulated Genes by Chromatin Immunoprecipitation.

HepG2 human hepatocellular carcinoma cell line was fixed using formaldehyde, resuspended in lysis buffer, and sonicated to shear chromatin. HMGB1/HMG-1/DNA complexes were immunoprecipitated using 5 µg Mouse Anti-Human HMGB1/HMG-1 PE-conjugated Monoclonal Antibody (Catalog # MAB1690) or control antibody (Catalog # MAB004) for 15 minutes in an ultrasonic bath, followed by Biotinylated Anti-Mouse IgG Secondary Antibody (Catalog # BAF007). Immunocomplexes were captured using 50 µL of MagCellect Streptavidin Ferrofluid (Catalog # MAG999) and DNA was purified using chelating resin solution. TheAFPpromoter was detected by standard PCR.
Detection of HMGB1 antibody in HCT-116 Human Cell Line antibody by Flow Cytometry.

Detection of HMGB1 in HCT‑116 Human Cell Line by Flow Cytometry.

HCT-116 human colorectal carcinoma cell line was stained with Mouse Anti-Human HMGB1/HMG-1 Monoclonal Antibody (Catalog # MAB1690, filled histogram) or isotype control antibody (Catalog # MAB0041, open histogram), followed by Phycoerythrin-conjugated Anti-Mouse IgG F(ab')2Secondary Antibody (Catalog # F0102B). To facilitate intracellular staining, cells were fixed with Flow Cytometry Fixation Buffer (Catalog # FC004) and permeabilized with Flow Cytometry Permeabilization/Wash Buffer I (Catalog # FC005). View our protocol for Staining Intracellular Molecules.
HMGB1/HMG-1 antibody in CCD-18Co Human Cell Line by Immunocytochemistry (ICC).

HMGB1/HMG‑1 in CCD‑18Co Human Cell Line.

HMGB1/HMG-1 was detected in immersion fixed CCD-18Co human colonic fibroblast cell line using Mouse Anti-Human HMGB1/HMG-1 Monoclonal Antibody (Catalog # MAB1690) at 10 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Mouse IgG Secondary Antibody (red, upper panel; Catalog # NL007) and counterstained with DAPI (blue, lower panel). Specific staining was localized to nuclei. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.
HMGB1/HMG-1 antibody in Human Breast Cancer Tissue by Immunohistochemistry (IHC-P).

HMGB1/HMG-1 in Human Breast Cancer Tissue.

HMGB1/HMG-1 was detected in immersion fixed paraffin-embedded sections of human breast cancer tissue using Mouse Anti-Human HMGB1/HMG-1 Monoclonal Antibody (Catalog # MAB1690) at 5 µg/mL overnight at 4 °C. Before incubation with the primary antibody, tissue was subjected to heat-induced epitope retrieval using Antigen Retrieval Reagent-Basic (Catalog # CTS013). Tissue was stained using the Anti-Mouse IgG VisUCyte™ HRP Polymer Antibody (brown; Catalog # VC001) and counterstained with hematoxylin (blue). Specific staining was localized to cell nuclei. View our protocol for IHC Staining with VisUCyte HRP Polymer Detection Reagents.
Detection of Rat HMGB1/HMG-1 by Western Blot

Detection of Rat HMGB1/HMG-1 by Western Blot

The effect of corticosterone on high-mobility group box-1 (HMGB1) release in primary cultured cortical astrocytes. (A) Increasing concentrations of corticosterone induced HMGB1 release from astrocytes. Cells were treated with the indicated concentrations of corticosterone for 24 h, and the amount of HMGB1 in the media was analyzed by Western blotting. Representative blots are shown (HMGB1: 30 kDa). The data are expressed as the mean ± SEM. * p < 0.05, ** p < 0.01 vs. basal (concentration “0”) (Dunnett’s test; n = 7–15). (B) Astrocytes were treated with 1 μM corticosterone for the indicated periods of time and the amount of HMGB1 in the media was analyzed by Western blotting. Representative blots are shown (HMGB1: 30 kDa). The data are expressed as the mean ± SEM. * p < 0.05, ** p < 0.01 vs. basal (time “0”) (Dunnett’s test; n = 3–15). (C) Astrocytes were treated with 1 μM corticosterone for 24 h, and media concentrations of HMGB1 were analyzed by ELISA. The data are expressed as the mean ± SEM. * p < 0.05 vs. vehicle (t-test; n = 5). Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/32344830), licensed under a CC-BY license. Not internally tested by R&D Systems.
Detection of Rat HMGB1/HMG-1 by Western Blot

Detection of Rat HMGB1/HMG-1 by Western Blot

The effect of corticosterone on high-mobility group box-1 (HMGB1) release in primary cultured cortical astrocytes. (A) Increasing concentrations of corticosterone induced HMGB1 release from astrocytes. Cells were treated with the indicated concentrations of corticosterone for 24 h, and the amount of HMGB1 in the media was analyzed by Western blotting. Representative blots are shown (HMGB1: 30 kDa). The data are expressed as the mean ± SEM. * p < 0.05, ** p < 0.01 vs. basal (concentration “0”) (Dunnett’s test; n = 7–15). (B) Astrocytes were treated with 1 μM corticosterone for the indicated periods of time and the amount of HMGB1 in the media was analyzed by Western blotting. Representative blots are shown (HMGB1: 30 kDa). The data are expressed as the mean ± SEM. * p < 0.05, ** p < 0.01 vs. basal (time “0”) (Dunnett’s test; n = 3–15). (C) Astrocytes were treated with 1 μM corticosterone for 24 h, and media concentrations of HMGB1 were analyzed by ELISA. The data are expressed as the mean ± SEM. * p < 0.05 vs. vehicle (t-test; n = 5). Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/32344830), licensed under a CC-BY license. Not internally tested by R&D Systems.
Detection of Human HMGB1/HMG-1 by Immunohistochemistry

Detection of Human HMGB1/HMG-1 by Immunohistochemistry

RAGE and HMGB1 are expressed in myofibroblasts of endarterectomised chronic thromboembolic tissues of CTEPH patients.One representative patient is shown (12 out of 15 patients (80%) displayed analogous staining patterns. Photograph showing the macroscopic aspect of a representative PEA specimen (A). Scale bar: 6 cm. Immunohistochemical expression of RAGE (B, scale bar: 20 µm), HMGB1 (C), vimentin (D), alpha-smooth muscle actin (E) and CD34 (F) on adjacent tissue sections of the PEA specimen shown in (A). Scale bar: 40 µm. RAGE receptor for advanced glycation endproducts, HMGB1 high mobility group box 1, CTEPH chronic thromboembolic pulmonary hypertension, PEA pulmonary endarterectomy. Image collected and cropped by CiteAb from the following publication (https://dx.plos.org/10.1371/journal.pone.0106440), licensed under a CC-BY license. Not internally tested by R&D Systems.

Applications for Human HMGB1/HMG‑1 Antibody

Application
Recommended Usage

Chromatin Immunoprecipitation (ChIP)

5 µg/5 x 106 cells
Sample: HepG2 human hepatocellular carcinoma cell line chromatin, AFP promoter detected by standard PCR

CyTOF-ready

Ready to be labeled using established conjugation methods. No BSA or other carrier proteins that could interfere with conjugation.

Immunocytochemistry

8-25 µg/mL
Sample: Immersion-fixed CCD-18Co human colonic fibroblast cell line and MCF-7 human breast cancer cell line

Immunohistochemistry

5-25 µg/mL
Sample: Immersion fixed paraffin-embedded sections of human breast cancer tissue

Intracellular Staining by Flow Cytometry

2.5 µg/106 cells
Sample: HCT‑116 human colorectal carcinoma cell line fixed with Flow Cytometry Fixation Buffer (Catalog # FC004) and permeabilized with Flow Cytometry Permeabilization/Wash Buffer I (Catalog # FC005)

Western Blot

0.05 µg/mL
Sample: HepG2 human hepatocellular carcinoma cell line, Jurkat human acute T cell leukemia cell line, and HEK293 human embryonic kidney cell line

Reviewed Applications

Read 3 reviews rated 4.3 using MAB1690 in the following applications:

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Flow Cytometry

Formulation, Preparation, and Storage

Purification

Protein A or G purified from hybridoma culture supernatant

Reconstitution

Reconstitute at 0.5 mg/mL in sterile PBS. For liquid material, refer to CoA for concentration.

Formulation

Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. *Small pack size (SP) is supplied either lyophilized or as a 0.2 µm filtered solution in PBS.

Shipping

Lyophilized product is shipped at ambient temperature. Liquid small pack size (-SP) is shipped with polar packs. Upon receipt, store immediately at the temperature recommended below.

Stability & Storage

Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 12 months from date of receipt, -20 to -70 °C as supplied.
  • 1 month, 2 to 8 °C under sterile conditions after reconstitution.
  • 6 months, -20 to -70 °C under sterile conditions after reconstitution.

Calculators

The reconstitution calculator allows you to quickly calculate the volume of a reagent to reconstitute your vial. Simply enter the mass of reagent and the target concentration and the calculator will determine the rest.

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Background: HMGB1/HMG-1

Human High-mobility group box 1 protein (HMGB1), previously known as HMG-1 or amphoterin, is a member of the high mobility group box family of non-histone chromosomal proteins (1‑3). Human HMGB1 is expressed as a 30 kDa, 215 amino acid (aa) single chain polypeptide containing three domains: two N-terminal globular, 70 aa positively charged DNA-binding domains (HMG boxes A and B), and a negatively charged 30 aa C-terminal region that contains only Asp and Glu (4, 5). Residues 27‑43 and 178‑184 contain a NLS. Posttranslational modifications of the molecule have been reported, with acetylation occurring on as many as 17 lysine residues (6). HMGB1 is expressed at high levels in almost all cells (2, 4). It was originally discovered as a nuclear protein that could bend DNA. Such bending stabilizes nucleosome formation and regulates the expression of select genes upon recruitment by DNA binding proteins (1, 7, 8). It is now known that HMGB1 can also act extracellularly, both as an inflammatory mediator that promotes monocyte migration and cytokine secretion, and as a mediator of T cell-dendritic cell interaction (1, 4, 7, 9, 10). The cytokine activity of HBMG1 is restricted to the HMG B box, (3) while the A box is associated with the helix-loop-helix domain of transcription factors (11). HMBG1 is released in response to cell death and as a secretion product. Although HMBG-1 does not possess a classic signal sequence, it appears to be secreted as an acetylated form via secretory endolysosome exocytosis (6, 12). Once secreted, HMGB1 transduces cellular signals through its high affinity receptor, RAGE and, possibly, TLR2 and TLR4 (1, 3, 4). Human HMGB1 is 100% aa identical to canine HMGB1 and 99% aa identical to mouse, rat, bovine and porcine HMGB1, respectively.

References

  1. Lotze, M.T. and K.J. Tracey (2005) Nat. Rev. Immunol. 5:331.
  2. Yang, H. et al. (2005) J. Leukoc. Biol. 78:1.
  3. Dumitriu, I.E. et al. (2005) Trends Immunol. 26:381.
  4. Degryse, B. and M. de Virgilio (2003) FEBS Lett. 553:11.
  5. Wen, L. et al. (1989) Nucleic Acids Res. 17:1197.
  6. Bonaldi, T. et al. (2003) EMBO J. 22:5551.
  7. Muller, S. et al. (2001) EMBO J. 20:4337.
  8. Bustin, M. (1999) Mol. Cell. Biol. 19:5237.
  9. Wang, H. et al. (1999) Science. 285:248.
  10. Dimitriu, I.E. et al. (2005) J. Immunol. 174:7506.
  11. Najima, Y. et al. (2005) J. Biol. Chem. 280:27523.
  12. Gardella, S. et al. (2002) EMBO Rep. 3:995.

Long Name

High Mobility Group Protein 1

Alternate Names

HMG-1, HMG1

Entrez Gene IDs

3146 (Human)

Gene Symbol

HMGB1

UniProt

P09429

Additional HMGB1/HMG-1 Products

Product Documents for Human HMGB1/HMG‑1 Antibody

Certificate of Analysis

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Product Specific Notices for Human HMGB1/HMG‑1 Antibody

For research use only

Citations for Human HMGB1/HMG‑1 Antibody

Customer Reviews for Human HMGB1/HMG‑1 Antibody (3)

4.3 out of 5
3 Customer Ratings
5 Stars
67%
4 Stars
0%
3 Stars
33%
2 Stars
0%
1 Stars
0%

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Customer Images

Showing  1 - 3 of 3 reviews Showing All
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  • Human HMGB1/HMG-1 Antibody
    Name: Anonymous
    Application: Western Blot
    Sample Tested: Cancer cell lysates
    Species: Human
    Verified Customer | Posted 10/17/2018
    Human HMGB1/HMG‑1 Antibody MAB1690
  • Human HMGB1/HMG-1 Antibody
    Name: Anonymous
    Application: Immunocytochemistry/Immunofluorescence
    Sample Tested: fibrotic lung
    Species: Mouse
    Verified Customer | Posted 10/13/2016
    Human HMGB1/HMG‑1 Antibody MAB1690
  • Human HMGB1/HMG‑1 Antibody
    Name: jinyan yu
    Application: Western Blot
    Sample Tested: Cell lysate
    Species: Human
    Verified Customer | Posted 01/23/2016
    I should say this is a good product. I purchased the small size, detected HMGB1 expression after TNF-a challenged. It is helpful.

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Protocols

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