WB result of Phospho-IRE1 (S724) Recombinant Rabbit mAb
Blocking/Diluting buffer and concentration: 5% NFDM/TBST
Primary antibody: Phospho-IRE1 (S724) Recombinant Rabbit mAb at 1/1000 dilution
Lane 1: untreated HeLa whole cell lysate 20 µg
Lane 2: Serum-starved HeLa treated with 100 nM Calyculin A for 30 minutes whole cell lysate 20 µg
Secondary antibody: Goat Anti- rabbit IgG, (H+L), HRP conjugated at 1/10000 dilution
Predicted MW: 110 kDa
Observed MW: 120 kDa
Phospho-IRE1 (S724) Recombinant Rabbit mAb (S-3480-15)
Phospho-IRE1 (S724) Recombinant Rabbit mAb (S-3480-15)
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Product Details
Product Details
Product Specification
| Host | Rabbit |
| Antigen | Phospho-IRE1 (S724) |
| Synonyms | Serine/threonine-protein kinase/endoribonuclease IRE1; Endoplasmic reticulum-to-nucleus signaling 1; Inositol-requiring protein 1 (hIRE1p); Ire1-alpha (IRE1a); IRE1; ERN1 |
| Immunogen | Synthetic Peptide |
| Location | Nucleus |
| Accession | O75460 |
| Clone Number | S-3480-15 |
| Antibody Type | Recombinant mAb |
| Isotype | IgG |
| Application | WB, ICC |
| Reactivity | Hu, Ms |
| Positive Sample | Serum-starved and Calyculin A treated HeLa, Calyculin A treated NIH/3T3 |
| Purification | Protein A |
| Concentration | 0.5 mg/ml |
| Conjugation | Unconjugated |
| Physical Appearance | Liquid |
| Storage Buffer | PBS, 40% Glycerol, 0.05% BSA, 0.03% Proclin 300 |
| Stability & Storage | 12 months from date of receipt / reconstitution, -20 °C as supplied |
Dilution
| application | dilution | species |
| Dot Blot | 1:1000 | |
| WB | 1:1000 | Hu, Ms |
| ICC | 1:100 | Hu |
Background
Phospho-IRE1 (S724) refers to the phosphorylated form of inositol-requiring enzyme 1-alpha (IRE1α) at serine residue 724, a critical post-translational modification that serves as a hallmark of IRE1α activation during the unfolded protein response (UPR). As an endoplasmic reticulum (ER) transmembrane protein kinase and endoribonuclease, IRE1α remains in an inactive, monomeric state under basal conditions through binding to the ER chaperone BiP/GRP78; however, upon accumulation of misfolded proteins within the ER lumen, dissociation from BiP triggers oligomerization and trans-autophosphorylation at multiple sites including S724 within its kinase activation loop. This phosphorylation event induces a conformational shift that licenses the RNase domain to excise a 26-nucleotide intron from X-box binding protein 1 (XBP1) mRNA, yielding the potent transcription factor XBP1s that upregulates ER-associated degradation (ERAD) machinery, chaperone expression, and phospholipid synthesis to restore proteostasis. Beyond XBP1 splicing, phospho-IRE1 (S724) also regulates the selective degradation of specific mRNAs and microRNAs through regulated IRE1-dependent decay (RIDD), modulating cellular outcomes ranging from survival to apoptosis depending on stress intensity and duration. Dysregulation of IRE1α phosphorylation at this site has been implicated in diverse pathologies including multiple myeloma, diabetes, neurodegenerative disorders, and inflammatory conditions, making S724-phosphorylated IRE1 a valuable biomarker for ER stress levels and an attractive therapeutic target for small molecule kinase inhibitors aimed at modulating the UPR signaling cascade.
Picture
Picture
Western Blot
WB result of Phospho-IRE1 (S724) Recombinant Rabbit mAb
Blocking/Diluting buffer and concentration: 5% NFDM/TBST
Primary antibody: Phospho-IRE1 (S724) Recombinant Rabbit mAb at 1/1000 dilution
Lane 1: untreated NIH/3T3 whole cell lysate 20 µg
Lane 2: NIH/3T3 treated with 100 nM Calyculin A for 30 minutes whole cell lysate 20 µg
Secondary antibody: Goat Anti- rabbit IgG, (H+L), HRP conjugated at 1/10000 dilution
Predicted MW: 110 kDa
Observed MW: 120 kDa
Dot Blot
Dot blot result of Phospho-IRE1 (S724) Recombinant Rabbit mAb
Lane 1: IRE1 (S724) phospho peptide
Lane 2: IRE1 (S724) unmodified peptide
Primary antibody: Phospho-IRE1 (S724) Recombinant Rabbit mAb at 1/1000 dilution
Secondary antibody: Goat Anti-rabbit IgG, (H+L), HRP conjugated at 1/10000 dilution
Immunocytochemistry
ICC analysis of HeLa cells starved for 3 hours then treated with 100nM Calyculin A for 30 minutes (top panel) and untreated HeLa cells (below panel). Anti- Phospho-IRE1 (S724) antibody was used at 1/100 dilution (Green) and incubated overnight at 4°C. Goat polyclonal Antibody to Rabbit IgG - H&L (Alexa Fluor® 488) was used as secondary antibody at 1/1000 dilution. The cells were fixed with 100% ice-cold methanol and permeabilized with 0.1% PBS-Triton X-100. Nuclei were counterstained with DAPI (Blue). Counterstain with tubulin (Red).
