Product: HMGB1 Antibody
Catalog: AF7020
Description: Rabbit polyclonal antibody to HMGB1
Application: WB IHC IF/ICC
Reactivity: Human, Mouse, Rat
Prediction: Pig, Bovine, Horse, Rabbit, Dog
Mol.Wt.: 20~25kD; 25kD(Calculated).
Uniprot: P09429
RRID: AB_2835324

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Product Info

WB 1:500-1:2000, IHC 1:50-1:200, IF/ICC 1:100-1:500
*The optimal dilutions should be determined by the end user.

WB: For western blot detection of denatured protein samples. IHC: For immunohistochemical detection of paraffin sections (IHC-p) or frozen sections (IHC-f) of tissue samples. IF/ICC: For immunofluorescence detection of cell samples. ELISA(peptide): For ELISA detection of antigenic peptide.

Pig(100%), Bovine(100%), Horse(100%), Rabbit(100%), Dog(100%)
HMGB1 Antibody detects endogenous levels of total HMGB1.
Cite Format: Affinity Biosciences Cat# AF7020, RRID:AB_2835324.
The antiserum was purified by peptide affinity chromatography using SulfoLink™ Coupling Resin (Thermo Fisher Scientific).
Rabbit IgG in phosphate buffered saline , pH 7.4, 150mM NaCl, 0.02% sodium azide and 50% glycerol. Store at -20 °C. Stable for 12 months from date of receipt.


Amphoterin; Chromosomal protein, nonhistone, HMG1; DKFZp686A04236; High mobility group 1; High mobility group box 1; High mobility group protein 1; High mobility group protein B1; high-mobility group (nonhistone chromosomal) protein 1; HMG-1; HMG1; HMG3; HMGB 1; HMGB1; HMGB1_HUMAN; NONHISTONE CHROMOSOMAL PROTEIN HMG1; SBP 1; Sulfoglucuronyl carbohydrate binding protein;



Ubiquituous. Expressed in platelets (PubMed:11154118).

High mobility group (HMG) proteins 1 and 2 are ubiquitous non-histone components of chromatin. Evidence suggests that the binding of HMG proteins to DNA induces alterations in the DNA architecture including DNA bending and unwinding of the helix. HMG proteins synergize with Oct-2, members of the NFκB family, ATF-2 and c-Jun to activate transcription.



Score>80(red) has high confidence and is suggested to be used for WB detection. *The prediction model is mainly based on the alignment of immunogen sequences, the results are for reference only, not as the basis of quality assurance.

Model Confidence:
High(score>80) Medium(80>score>50) Low(score<50) No confidence

PTMs - P09429 As Substrate

Site PTM Type Enzyme
K3 Acetylation
K3 Methylation
K12 Acetylation
K12 Ubiquitination
S15 Phosphorylation
Y16 Phosphorylation
C23 S-Nitrosylation
R24 Methylation
K30 Acetylation
K30 Ubiquitination
S35 Phosphorylation
S39 Phosphorylation
S42 Phosphorylation
K43 Acetylation
K43 Methylation
K43 Ubiquitination
K44 Acetylation
K44 Ubiquitination
S46 Phosphorylation
K50 Ubiquitination
T51 Phosphorylation
S53 Phosphorylation
K55 Acetylation
K55 Ubiquitination
K59 Acetylation
K59 Methylation
K59 Ubiquitination
K65 Acetylation
K65 Ubiquitination
Y71 Phosphorylation
K76 Acetylation
K76 Ubiquitination
T77 Phosphorylation
Y78 Phosphorylation
K82 Acetylation
K82 Ubiquitination
K90 Acetylation
K90 Ubiquitination
S100 Phosphorylation
C106 S-Nitrosylation
S107 Phosphorylation
Y109 Phosphorylation
K112 Methylation
K112 Ubiquitination
K114 Methylation
K114 Sumoylation
K114 Ubiquitination
S121 Phosphorylation
K127 Ubiquitination
K128 Acetylation
K128 Ubiquitination
T136 Phosphorylation
K141 Ubiquitination
Y144 Phosphorylation
K146 Ubiquitination
K147 Ubiquitination
K154 Acetylation
Y155 Phosphorylation
K157 Acetylation
K157 Ubiquitination
Y162 Phosphorylation
K173 Acetylation
K177 Acetylation
K177 Sumoylation
S181 Phosphorylation

Research Backgrounds


Multifunctional redox sensitive protein with various roles in different cellular compartments. In the nucleus is one of the major chromatin-associated non-histone proteins and acts as a DNA chaperone involved in replication, transcription, chromatin remodeling, V(D)J recombination, DNA repair and genome stability. Proposed to be an universal biosensor for nucleic acids. Promotes host inflammatory response to sterile and infectious signals and is involved in the coordination and integration of innate and adaptive immune responses. In the cytoplasm functions as sensor and/or chaperone for immunogenic nucleic acids implicating the activation of TLR9-mediated immune responses, and mediates autophagy. Acts as danger associated molecular pattern (DAMP) molecule that amplifies immune responses during tissue injury. Released to the extracellular environment can bind DNA, nucleosomes, IL-1 beta, CXCL12, AGER isoform 2/sRAGE, lipopolysaccharide (LPS) and lipoteichoic acid (LTA), and activates cells through engagement of multiple surface receptors. In the extracellular compartment fully reduced HMGB1 (released by necrosis) acts as a chemokine, disulfide HMGB1 (actively secreted) as a cytokine, and sulfonyl HMGB1 (released from apoptotic cells) promotes immunological tolerance. Has proangiogdenic activity (By similarity). May be involved in platelet activation (By similarity). Binds to phosphatidylserine and phosphatidylethanolamide (By similarity). Bound to RAGE mediates signaling for neuronal outgrowth (By similarity). May play a role in accumulation of expanded polyglutamine (polyQ) proteins such as huntingtin (HTT) or TBP.

Nuclear functions are attributed to fully reduced HGMB1. Associates with chromatin and binds DNA with a preference to non-canonical DNA structures such as single-stranded DNA, DNA-containing cruciforms or bent structures, supercoiled DNA and ZDNA. Can bent DNA and enhance DNA flexibility by looping thus providing a mechanism to promote activities on various gene promoters by enhancing transcription factor binding and/or bringing distant regulatory sequences into close proximity. May have an enhancing role in nucleotide excision repair (NER) (By similarity). However, effects in NER using in vitro systems have been reported conflictingly. May be involved in mismatch repair (MMR) and base excision repair (BER) pathways. May be involved in double strand break repair such as non-homologous end joining (NHEJ) (By similarity). Involved in V(D)J recombination by acting as a cofactor of the RAG complex: acts by stimulating cleavage and RAG protein binding at the 23 bp spacer of conserved recombination signal sequences (RSS) (By similarity). In vitro can displace histone H1 from highly bent DNA (By similarity). Can restructure the canonical nucleosome leading to relaxation of structural constraints for transcription factor-binding (By similarity). Enhances binding of sterol regulatory element-binding proteins (SREBPs) such as SREBF1 to their cognate DNA sequences and increases their transcriptional activities (By similarity). Facilitates binding of TP53 to DNA. Proposed to be involved in mitochondrial quality control and autophagy in a transcription-dependent fashion implicating HSPB1; however, this function has been questioned (By similarity). Can modulate the activity of the telomerase complex and may be involved in telomere maintenance (By similarity).

In the cytoplasm proposed to dissociate the BECN1:BCL2 complex via competitive interaction with BECN1 leading to autophagy activation. Involved in oxidative stress-mediated autophagy. Can protect BECN1 and ATG5 from calpain-mediated cleavage and thus proposed to control their proautophagic and proapoptotic functions and to regulate the extent and severity of inflammation-associated cellular injury (By similarity). In myeloid cells has a protective role against endotoxemia and bacterial infection by promoting autophagy (By similarity). Involved in endosomal translocation and activation of TLR9 in response to CpG-DNA in macrophages (By similarity).

In the extracellular compartment (following either active secretion or passive release) involved in regulation of the inflammatory response. Fully reduced HGMB1 (which subsequently gets oxidized after release) in association with CXCL12 mediates the recruitment of inflammatory cells during the initial phase of tissue injury; the CXCL12:HMGB1 complex triggers CXCR4 homodimerization. Induces the migration of monocyte-derived immature dendritic cells and seems to regulate adhesive and migratory functions of neutrophils implicating AGER/RAGE and ITGAM (By similarity). Can bind to various types of DNA and RNA including microbial unmethylated CpG-DNA to enhance the innate immune response to nucleic acids. Proposed to act in promiscuous DNA/RNA sensing which cooperates with subsequent discriminative sensing by specific pattern recognition receptors (By similarity). Promotes extracellular DNA-induced AIM2 inflammasome activation implicating AGER/RAGE. Disulfide HMGB1 binds to transmembrane receptors, such as AGER/RAGE, TLR2, TLR4 and probably TREM1, thus activating their signal transduction pathways. Mediates the release of cytokines/chemokines such as TNF, IL-1, IL-6, IL-8, CCL2, CCL3, CCL4 and CXCL10. Promotes secretion of interferon-gamma by macrophage-stimulated natural killer (NK) cells in concert with other cytokines like IL-2 or IL-12. TLR4 is proposed to be the primary receptor promoting macrophage activation and signaling through TLR4 seems to implicate LY96/MD-2. In bacterial LPS- or LTA-mediated inflammatory responses binds to the endotoxins and transfers them to CD14 for signaling to the respective TLR4:LY96 and TLR2 complexes. Contributes to tumor proliferation by association with ACER/RAGE (By similarity). Can bind to IL1-beta and signals through the IL1R1:IL1RAP receptor complex. Binding to class A CpG activates cytokine production in plasmacytoid dendritic cells implicating TLR9, MYD88 and AGER/RAGE and can activate autoreactive B cells. Via HMGB1-containing chromatin immune complexes may also promote B cell responses to endogenous TLR9 ligands through a B-cell receptor (BCR)-dependent and ACER/RAGE-independent mechanism (By similarity). Inhibits phagocytosis of apoptotic cells by macrophages; the function is dependent on poly-ADP-ribosylation and involves binding to phosphatidylserine on the cell surface of apoptotic cells (By similarity). In adaptive immunity may be involved in enhancing immunity through activation of effector T cells and suppression of regulatory T (TReg) cells. In contrast, without implicating effector or regulatory T-cells, required for tumor infiltration and activation of T-cells expressing the lymphotoxin LTA:LTB heterotrimer thus promoting tumor malignant progression (By similarity). Also reported to limit proliferation of T-cells (By similarity). Released HMGB1:nucleosome complexes formed during apoptosis can signal through TLR2 to induce cytokine production. Involved in induction of immunological tolerance by apoptotic cells; its pro-inflammatory activities when released by apoptotic cells are neutralized by reactive oxygen species (ROS)-dependent oxidation specifically on Cys-106. During macrophage activation by activated lymphocyte-derived self apoptotic DNA (ALD-DNA) promotes recruitment of ALD-DNA to endosomes (By similarity).


Phosphorylated at serine residues. Phosphorylation in both NLS regions is required for cytoplasmic translocation followed by secretion.

Acetylated on multiple sites upon stimulation with LPS. Acetylation on lysine residues in the nuclear localization signals (NLS 1 and NLS 2) leads to cytoplasmic localization and subsequent secretion (By similarity). Acetylation on Lys-3 results in preferential binding to DNA ends and impairs DNA bending activity (By similarity).

Reduction/oxidation of cysteine residues Cys-23, Cys-45 and Cys-106 and a possible intramolecular disulfide bond involving Cys-23 and Cys-45 give rise to different redox forms with specific functional activities in various cellular compartments: 1- fully reduced HMGB1 (HMGB1C23hC45hC106h), 2- disulfide HMGB1 (HMGB1C23-C45C106h) and 3- sulfonyl HMGB1 (HMGB1C23soC45soC106so).

Poly-ADP-ribosylated by PARP1 when secreted following stimulation with LPS (By similarity).

In vitro cleavage by CASP1 is liberating a HMG box 1-containing peptide which may mediate immunogenic activity; the peptide antagonizes apoptosis-induced immune tolerance. Can be proteolytically cleaved by a thrombin:thrombomodulin complex; reduces binding to heparin and proinflammatory activities (By similarity).

Subcellular Location:

Nucleus. Chromosome. Cytoplasm. Secreted. Cell membrane>Peripheral membrane protein>Extracellular side. Endosome. Endoplasmic reticulum-Golgi intermediate compartment.
Note: In basal state predominantly nuclear. Shuttles between the cytoplasm and the nucleus (PubMed:12231511, PubMed:17114460). Translocates from the nucleus to the cytoplasm upon autophagy stimulation (PubMed:20819940). Release from macrophages in the extracellular milieu requires the activation of NLRC4 or NLRP3 inflammasomes (By similarity). Passively released to the extracellular milieu from necrotic cells by diffusion, involving the fully reduced HGMB1 which subsequently gets oxidized (PubMed:19811284). Also released from apoptotic cells (PubMed:16855214, PubMed:18631454). Active secretion from a variety of immune and non-immune cells such as macrophages, monocytes, neutrophils, dendritic cells and natural killer cells in response to various stimuli such as LPS and cytokines involves a nonconventional secretory process via secretory lysosomes (PubMed:12231511, PubMed:14532127, PubMed:15944249). Secreted by plasma cells in response to LPS (By similarity). Found on the surface of activated platelets (PubMed:11154118). An increased chromatin association is observed when associated with the adenovirus protein pVII (PubMed:27362237).

Extracellular region or secreted Cytosol Plasma membrane Cytoskeleton Lysosome Endosome Peroxisome ER Golgi apparatus Nucleus Mitochondrion Manual annotation Automatic computational assertionSubcellular location
Tissue Specificity:

Ubiquituous. Expressed in platelets.

Subunit Structure:

Interacts (fully reduced HMGB1) with CXCL12; probably in a 1:2 ratio involving two molecules of CXCL12, each interacting with one HMG box of HMGB1; inhibited by glycyrrhizin. Associates with the TLR4:LY96 receptor complex. Component of the RAG complex composed of core components RAG1 and RAG2, and associated component HMGB1 or HMGB2 (By similarity). Interacts (in cytoplasm upon starvation) with BECN1; inhibits the interaction of BECN1 and BCL2 leading to promotion of autophagy. Interacts with KPNA1; involved in nuclear import. Interacts with SREBF1, TLR2, TLR4, TLR9, PTPRZ1, APEX1, FEN1, POLB, TERT (By similarity). Interacts with IL1B, AGER, MSH2, XPA, XPC, HNF1A, TP53. Interacts with CD24; the probable CD24:SIGLEC10 complex is proposed to inhibit HGMB1-mediated tissue damage immune response. Interacts with THBD; prevents HGMB1 interaction with ACER/RAGE and inhibits HGMB1 proinflammatory activity. Interacts with HAVCR2; impairs HMGB1 binding to B-DNA and likely HMGB1-mediated innate immune response (By similarity). Interacts with XPO1; mediating nuclear export (By similarity). Interacts with HTT (wild-type and mutant HTT with expanded polyglutamine repeat).

(Microbial infection) Interacts with adenovirus protein pVII; this interaction immobilizes HMGB1 on chromatin, thus preventing its release from cell and subsequent inflammation activation.


HMG box 2 mediates proinflammatory cytokine-stimulating activity and binding to TLR4 (PubMed:12765338, PubMed:20547845). However, not involved in mediating immunogenic activity in the context of apoptosis-induced immune tolerance (PubMed:24474694).

The acidic C-terminal domain forms a flexible structure which can reversibly interact intramolecularily with the HMG boxes and modulate binding to DNA and other proteins (PubMed:23063560).

Belongs to the HMGB family.

Research Fields

· Cellular Processes > Transport and catabolism > Autophagy - animal.   (View pathway)

· Cellular Processes > Cell growth and death > Necroptosis.   (View pathway)

· Genetic Information Processing > Replication and repair > Base excision repair.


1). Mitochondrial Disruption Nanosystem Simultaneously Depressed Programmed Death Ligand-1 and Transforming Growth Factor-β to Overcome Photodynamic Immunotherapy Resistance. ACS nano, 2024 (PubMed: 38227812) [IF=17.1]

2). Design of a self-driven probiotic-CRISPR/Cas9 nanosystem for sono-immunometabolic cancer therapy. Nature Communications, 2022 (PubMed: 36550159) [IF=16.6]

3). Lenvatinib- and vadimezan-loaded synthetic high-density lipoprotein for combinational immunochemotherapy of metastatic triple-negative breast cancer. Acta Pharmaceutica Sinica B, 2022 (PubMed: 36176911) [IF=14.5]

Application: IF/ICC    Species: Mouse    Sample: 4T1 cells

Figure 3. Uptake and efficacy of sHDLs. (A) Flow cytometry analysis of time-dependent cellular uptake of Cy5-labelled sHDL by 4T1 cells. (B) Confocal images of 4T1 cells treated with Cy5-labelled sHDL at different time points. The nucleus and β-actin were stained with DAPI and Actin-Tracker Green, respectively. Scale bar = 20 μm. (C) Cell viability of 4T1 cells after 48 h exposure to LEN, L-sHDL, and LV-sHDL of various concentrations. Quantification of extracellular ATP (D) and HMGB1 (E) in the medium of 4T1 cells after the treatment of different sHDLs. (F) Confocal images of 4T1 cells after 4 h treatment with different sHDLs and another 12 h incubation with medium. The nucleus, HMGB1, and CRT were stained with DAPI, anti-HMGB1 antibody-Alexa Fluor® 647, and anti-CRT antibody-Alexa Fluor® 488, respectively. Scale bar = 20 μm. (G) Quantification of IFN-α and IFN-β secreted by 4T1 cells after the treatment of different sHDLs. Statistical significance was calculated using a two-sided one-way ANOVA test. The data are presented as the mean ± SD (n = 3).

4). Treadmill exercise improves neurological function by inhibiting autophagy and the binding of HMGB1 to Beclin1 in MCAO juvenile rats. LIFE SCIENCES, 2020 (PubMed: 31926245) [IF=6.1]

Application: WB    Species: Rat    Sample:

Fig. 6. Effects of treadmill exercise and RAPA injection on HMGB1 and Beclin1 expression in the ischemic penumbra. (A) HMGB1/Beclin1-positive cells in the ischemic penumbra at 7 days after MCAO. Scale bar, 20 μm. (B) Histogram of the number of HMGB1/Beclin1-positive cells in the ischemic penumbra at 3 and 7 days after MCAO. (C) Beclin1 and cytoplasmic HMGB1 proteins in the ischemic penumbra at 3 and 7 days after MCAO. (D) Histogram of cytoplasmic HMGB1 protein in the ischemic penumbra at 3 and 7 days after MCAO. (E) Histogram of Beclin1 protein in the ischemic penumbra at 3 and 7 days after MCAO. *P < 0.05, **P < 0.01, and ***P < 0.001 versus the same period for the MCAO group. #P < 0.05 and ##P < 0.05 versus the same period for the EX + RAPA + MCAO group.

5). Nesfatin-1 regulates the HMGB1-TLR4-NF-κB signaling pathway to inhibit inflammation and its effects on the random skin flap survival in rats. International Immunopharmacology, 2023 (PubMed: 37633241) [IF=5.6]

6). Effect of paeoniflorin on distal survival of random flaps. International Immunopharmacology, 2022 (PubMed: 35093690) [IF=5.6]

7). GLP-1 mediates the neuroprotective action of crocin against cigarette smoking-induced cognitive disorders via suppressing HMGB1-RAGE/TLR4-NF-κB pathway. International Immunopharmacology, 2022 (PubMed: 35785730) [IF=5.6]

Application: WB    Species: Rat    Sample: hippocampal

Fig. 5. Effect of crocin on hippocampal HMGB-1 level and expression as well as RAGE expression in a rat model of CS-induced cognitive disorders. (A) HMGB1 level (pg/g tissue); n = 6, (B) western blot showing HMGB1 and RAGE expression, (C) bar graph reflecting HMGB1 expression, and (D) bar graph reflecting RAGE expression. Protein expression level was normalized to β-actin and shown relative to that of control; n = 3. Values were expressed as means ± SD. Significant difference were presented at *p < 0.05 and ***p < 0.001; using One-way ANOVA followed by Tukey's Honestly Significant Difference (HSD) post-hoc. C: control; Cro: crocin; CS: cigarettes smoke; HMGB1: High mobility group box 1 Glucagon-like peptide-1; RAGE: Receptor for advanced glycation end products.

8). Zuojin Pill attenuates Helicobacter pylori-induced chronic atrophic gastritis in rats and improves gastric epithelial cells function in GES-1 cells. Journal of Ethnopharmacology, 2022 (PubMed: 34808298) [IF=5.4]

9). Cardiomyocyte-derived HMGB1 takes a protective role in CVB3-induced viral myocarditis via inhibiting cardiac apoptosis. Immunology and Cell Biology, 2023 (PubMed: 37253434) [IF=4.0]

10). HMGB1/RAGE axis mediates the apoptosis, invasion, autophagy, and angiogenesis of the renal cell carcinoma. OncoTargets and Therapy, 2018 (PubMed: 30122942) [IF=4.0]

Application: WB    Species: human    Sample: RCC cell

Figure 1 |HMGB1 knockdown decreased the viability of A498 and ACHN cells.Notes: (A) The RAGE and HMGB1 proteins in RCC cell lines were detected by Western blot.

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