HMGB1 Antibody (YA361)

HMGB1 Antibody (YA361) is a Rabbit-derived and non-conjugated IgG monoclonal antibody, targeting to HMGB1.

HMGB1 is a multifunctional redox-sensitive protein with diverse roles in various cellular compartments. In the nucleus, it acts as a major chromatin-associated non-histone protein and serves as a DNA chaperone involved in replication, transcription, chromatin remodeling, V(D)J recombination, DNA repair, and genome stability. In the cytoplasm, it functions as a sensor and/or chaperone for immunogenic nucleic acids, implicating TLR9-mediated immune response activation and mediating autophagy. As a danger-associated molecular pattern (DAMP) molecule, HMGB1 amplifies immune responses during tissue injury. When released into the extracellular environment, it binds to DNA, nucleosomes, IL-1 beta, CXCL12, AGER isoform 2/sRAGE, lipopolysaccharide (LPS), and lipoteichoic acid (LTA), activating cells through multiple surface receptors. 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. It exhibits proangiogenic activity (by similar) and may be involved in platelet activation (by similar). It binds phosphatidylserine and phosphatidylethanolamide (by similar) and mediates neuronal outgrowth signaling via RAGE (by similar). It may also play a role in the accumulation of expanded polyglutamine (polyQ) proteins such as huntingtin (HTT) or TBP.

Free

P09429

Predicted band size: 25 kDa; Observed band size: 25 kDa

Protein A affinity purified.

Cytoplasm, Nucleus, Cell membrane, Secreted, Chromosome

Non-conjugated

Unmodified

AB_3102332

Epigenetics and Nuclear Signaling

Primary Antibody; Recombinant Rabbit Monoclonal Antibody

Recombinant, Monoclonal

Rabbit

Human, Mouse, Rat

WB: 1:500-1:5000; ICC/IF: 1:100; IHC-P: 1:5000; FC: 1:50

• 
  Western blot analysis of extracts from Hela(lane 2(20μg), C6 (lane 3(20μg) and HepG2(lane 4(20μg) using HMGB1 (HY-P80183) Rabbit mAb. Proteins were transferred to a PVDF membrane and blocked with 5% non-fat milk in TBST for 2 hour at room temperature. The primary antibody (1/1000) and Loading control antibody (Beta Actin, HY-P80438, 1/10000) was used in 5% non-fat milk in TBST at 4°C overnight. Goat Anti-Mouse/Rabbit IgG-HRP Secondary Antibody (1/10000) was used for 1 hour at room temperature.
• 
  Immunocytochemistry analysis of HELA cells labeling HMGB1 with HMGB1 Antibody (HY-P80183) at 1/50 dilution. Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 for 10 minutes at room temperature, then blocked with QuickBlock™ Blocking Buffer for Immunol Staining for 10 min at room temperature. Cells were then incubated with HMGB1 Antibody (HY-P80183) at 1/100 dilution in QuickBlock™ Blocking Buffer for Immunol Staining at 4 ℃. Alexa Fluor® 488-conjugated AffiniPure Goat Anti-Rabbit IgG H&L（HY-P8002, Green) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. The Nuclear counterstain was DAPI (Blue).
• 
  Immunocytochemistry analysis of HELA cells labeling HMGB1 with HMGB1 Antibody (HY-P80183) at 1/100 dilution. Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 for 10 minutes at room temperature, then blocked with QuickBlock™ Blocking Buffer for Immunol Staining for 10 min at room temperature. Cells were then incubated with HMGB1 Antibody (HY-P80183) at 1/100 dilution in QuickBlock™ Blocking Buffer for Immunol Staining at 4 ℃. Alexa Fluor® 488-conjugated AffiniPure Goat Anti-Rabbit IgG H&L（HY-P8002, Green) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. The Nuclear counterstain was DAPI (Blue).
• 
  Immunohistochemical analysis of paraffin-embedded Mouse brain tissue using HMGB1 Antibody. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) for 8 minutes. The tissues were blocked in QuickBlock for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HY-P80183, 1/5000) in 4℃ overnight. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
• 
  Immunohistochemical analysis of paraffin-embedded Mouse brain tissue using HMGB1 Antibody. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) for 8 minutes. The tissues were blocked in QuickBlock for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HY-P80183, 1/5000) in 4℃ overnight. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
• 
  Flow cytometric analysis of 1X10^6 Hela cells labeling HMGB1 Antibody (HY-P80183, red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. Then stained with the primary antibody at 1/50 dilution for an hour at 4℃. Alexa Fluor® 488-conjugated AffiniPure Goat Anti-Rabbit IgG H&L (HY-P8002) was used as the secondary antibody at 1/1,000 dilution for 30 minutes at 4℃. Rabbit IgG Isotype Control (HY-P80879, blue) was used as the isotype control, cells without incubation with primary antibody were used as the unlabeled control (black).

WB, ICC/IF, IHC-P, FC

Liquid

Supplied in 1*TBS (pH7.4), 0.05% BSA and 40% Glycerol. Preservative: 0.05% Sodium Azide.

Stored at -20°C for 1 year. Avoid repeated freeze / thaw cycles.

Shipping with blue ice.

IgG

Endogenous

Synthetic peptide corresponding to Human HMGB1.AA range:151-200.