Anti-phospho-IRE-1α (Ser724) Antibody, clone 1M2 ZooMAb® Rabbit Monoclonal
SIGMA/ZRB1072 - recombinant, expressed in HEK 293 cells
Synonym: Anti-IRE1; Anti-IRE1P; Anti-IRE1a; Anti-hIRE1p
Product Type: Chemical
Immunocytochemistry Enhanced Validation - Recombinant Antibody Technology Immunofluorescent analysis of C2C12 cells was performed using a 1:1,000 dilution of Cat. No. ZRB1072, Anti-phospho-IRE-1a (Ser724), clone 1M2 ZooMAb® Rabbit Monoclonal and visualized with a Goat Anti-Rabbit secondary antibody conjugated to Alexa Fluor® 488. Actin filaments have been labeled with phalloidin (Red). Nucleus is stained with DAPI (Blue). This antibody positively stains the endoplasmic reticulum.
Immunocytochemistry Enhanced Validation - Recombinant Antibody Technology Immunofluorescent analysis of C2C12 cells was performed using a 1:1,000 dilution of Cat. No. ZRB1072, Anti-phospho-IRE-1a (Ser724), clone 1M2 ZooMAb® Rabbit Monoclonal and visualized with a Goat Anti-Rabbit secondary antibody conjugated to Alexa Fluor® 488. Actin filaments have been labeled with phalloidin (Red). Nucleus is stained with DAPI (Blue). This antibody positively stains the endoplasmic reticulum.
Immunocytochemistry Enhanced Validation - Recombinant Antibody Technology Immunofluorescent analysis of C2C12 cells was performed using a 1:1,000 dilution of Cat. No. ZRB1072, Anti-phospho-IRE-1a (Ser724), clone 1M2 ZooMAb® Rabbit Monoclonal and visualized with a Goat Anti-Rabbit secondary antibody conjugated to Alexa Fluor® 488. Actin filaments have been labeled with phalloidin (Red). Nucleus is stained with DAPI (Blue). This antibody positively stains the endoplasmic reticulum.
Western Blotting Enhanced Validation - Recombinant Antibody Technology Lysates from untreated NIH3T3 (Lane 1) and NIH3T3 cells treated with Calyculin A (50 nM; 30 min) (Lane 2) were probed with Cat. No. ZRB1072, Anti-phospho-IRE-1a (Ser724), clone 1M2 ZooMAb® Rabbit Monoclonal (1:10,000 dilution). Proteins were visualized using a Donkey Anti-Rabbit IgG secondary antibody conjugated to HRP and a chemiluminescence detection system. Arrow indicates phospho-IRE-1a (Ser724) (~110 kDa).
Affinity Binding Assay Enhanced Validation - Recombinant Antibody Technology 100 mg/mL of non-specific control peptide (Run 1) or IRE-1a phospho Ser724 peptide (Run 2) were analyzed by affinity binding assay using Cat. No. ZRB1072, Anti-phospho-IRE-1α (Ser724), clone 1M2 ZooMAb® Rabbit Monoclonal. The binding of this antibody to IRE-1a phospho Ser724 peptide displayed a KD of 9.5 x 10-8 whereas binding to non-specific control peptide displayed a KD of 9.2 x 10-4.
Peptide Inhibition Assay Enhanced Validation - Recombinant Antibody Technology Target band detection in lysate from NIH3T3 cells treated with 50 nM Calyculin A was prevented by preblocking of a representative lot of Cat. No. ZRB1072, Anti-phospho-IRE-1α (Ser724), clone 1M2 ZooMAb® Rabbit Monoclonal with the immunogen phosphopeptide, but not the corresponding non-phosphopeptide. Lane 1: Anti-IRE-1a phospho Ser724, clone 1M2 tested without any peptide. Lane 2: Anti-IRE-1a phospho Ser724, clone 1M2 blocked with phosphorylated IRE-1a peptide. Lane 3: Anti-IRE-1a phospho Ser724, clone 1M2 blocked with non-phosphorylated IRE-1a peptide.
12 Principles of Green Chemistry: Principle 1—Waste Prevention. This product has a smaller environmental footprint through reduction of waste in the manufacturing process.
12 Principles of Green Chemistry: Principle 4—Designing Safer Chemicals
Chemical products should be designed to affect their desired function while minimizing their toxicity.
Chemical products should be designed to affect their desired function while minimizing their toxicity.
12 Principles of Green Chemistry: Principle 6—Design for Energy Efficiency. This product was re-engineered to utilize less energy in the manufacturing process.
| antibody form | purified antibody |
| antibody product type | primary antibodies |
| biological source | rabbit |
| clone | 1M2, recombinant monoclonal |
| conjugate | unconjugated |
| description | recombinant, expressed in HEK 293 cells |
| enhanced validation | recombinant expression Learn more about Antibody Enhanced Validation  |
| epitope sequence | C-terminal half |
| form | lyophilized |
| greener alternative category | Aligned |
| greener alternative product characteristics | Waste Prevention Designing Safer Chemicals Design for Energy Efficiency Learn more about the Principles of Green Chemistry . |
| isotype | IgG |
| mol wt | calculated mol wt 109.74 kDa |
| observed mol wt ~110 kDa | |
| packaging | antibody small pack of 25 μL |
| product line | ZooMAb® learn more |
| Protein ID accession no. | NP_001424.3?  |
| Quality Level | 200 |
| recombinant | expressed in HEK 293 cells |
| shipped in | ambient |
| species reactivity | human, mouse |
| species reactivity (predicted by homology) | rat |
| storage temp. | 2-8°C |
| technique(s) | affinity binding assay: suitable |
| immunocytochemistry: suitable | |
| western blot: suitable | |
| UniProt accession no. | O75460 |
| Application: | Quality Control Testing Evaluated by Western Blotting in lysate from HeLa cells treated with Calyculin A. Western Blotting Analysis: A 1:10,000 dilution of this antibody detected phospho-IRE- 1a (Ser724) lysate from NIH3T3 cells treated with Calyculin A (50 nM; 30 min.). Tested applications Western Blotting Analysis: A 1:10,000 dilution from a representative lot detected phospho-IRE-1a (Ser724) in untreated NIH3T3 and NIH3T3 treated with Calyculin A. Affinity Binding Assay: A representative lot of this antibody bound IRE-1a (phospho Ser724) peptide with a KD of 9.5 x 10-8 in an affinity binding assay. Immunocytochemistry Analysis: A 1:1,000 dilution from a representative lot detected phospho-IRE-1a (Ser724) in C2C12 cells.. Peptide Inhibition Assay: Target band detection in lysate from NIH3T3 cells treated with 50 nM Calyculin A was prevented by preblocking of a representative lot of phospho-IRE-1a (Ser724) with the immunogen phosphopeptide, but not the corresponding non-phosphopeptide. Note: Actual optimal working dilutions must be determined by end user as specimens, and experimental conditions may vary with the end user |
| Application: | Anti-phospho-IRE-1a (Ser724), clone 1M2 ZooMAb®, Cat. No. ZRB1072, is a Rabbit monoclonal antibody that targets phospho-IRE1a (Ser724) and is tested for use in Affinity Binding Assay, Immunocytochemistry, Peptide Inhibition Assay, and Western Blotting. |
| Disclaimer: | Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals. |
| General description: | We are committed to bringing you greener alternative products, which adhere to one or more of The 12 Principles of Green Chemistry.This antibody is Preservative-free, produced without the harm or sacrifice of animals and exceptionally stable to allow for ambient shipping and storage if needed and thus aligns with "Waste Prevention", "Designing Safer Chemicals" and "Design for Energy Efficiency". Click here for more information. |
| General description: | ZooMAb® antibodies represent an entirely new generation of recombinant monoclonal antibodies. Each ZooMAb® antibody is manufactured using our proprietary recombinant expression system, purified to homogeneity, and precisely dispensed to produce robust and highly reproducible lot-to-lot consistency. Only top-performing clones are released for use by researchers. Each antibody is validated for high specificity and affinity across multiple applications, including its most commonly used application. ZooMAb® antibodies are reliably available and ready to ship when you need them. Learn more about ZooMAb here. |
| Immunogen: | KLH-conjugated linear peptide corresponding to 10 amino acids surrounding phosphoserine 724 from the C-terminal, cytoplasmic domain of human Serine/threonine-protein kinase/endoribonuclease IRE1 (IRE-1a). |
| Legal Information: | ZooMAb is a registered trademark of Merck KGaA, Darmstadt, Germany |
| Physical form: | Purified recombinant rabbit monoclonal antibody IgG, lyophilized in PBS with 5% Trehalose, normal appearance a coarse or translucent resin. The PBS/trehalose components in the ZooMAb formulation can have the appearance of a semi-solid (bead like gel) after lyophilization. This is a normal phenomenon. Please follow the recommended reconstitution procedure in the data sheet to dissolve the semi-solid, bead-like, gel-appearing material. The resulting antibody solution is completely stable and functional as proven by full functional testing. Contains no biocide or preservatives, such as azide, or any animal by-products. Larger pack sizes provided as multiples of 25 μL. |
| Reconstitution: | 300 μg/mL after reconstitution at 25 μL per vial. Please refer to guidance on suggested starting dilutions and/or titers per application and sample type. |
| Specificity: | Clone IM2 is a ZooMAb® rabbit recombinant monoclonal antibody that specifically detects Serine/threonine-protein kinase/endoribonuclease IRE1 (IRE-1a) phosphorylated on serine 724. |
| Storage and Stability: | Recommend storage of lyophilized product at 2-8°C; Before reconstitution, micro-centrifuge vials briefly to spin down material to bottom of the vial; Reconstitute each vial by adding 25 μL of filtered lab grade water or PBS; Reconstituted antibodies can be stored at 2-8°C, or -20°C for long term storage. Avoid repeated freeze-thaws. |
| Target description: | Serine/threonine-protein kinase/endoribonuclease IRE1 (UniProt: O75460; also known as Endoplasmic reticulum-to-nucleus signaling 1, Inositol-requiring protein 1, hIRE1p, Ire1-alpha, IRE 1a ) is encoded by the ERN1 (also known a IRE1) gene (Gene ID: 2081) in human. IRE1a is a single-pass type I membrane protein that is synthesized with a signal peptide (aa 1-18), which is subsequently cleaved off to generate the mature form that contains a lumenal domain (aa 19-443), a transmembrane domain (aa 444-464), and a long cytoplasmic domain (aa 465-977). It serves as a serine/threonine-protein kinase and endoribonuclease that acts as a key sensor for the endoplasmic reticulum unfolded protein response (UPR). IRE1 activity can induce both protective UPR and apoptotic downstream signaling events. In unstressed cells, the endoplasmic reticulum luminal domain is maintained in its inactive monomeric state by binding to the endoplasmic reticulum chaperone HSPA5/BiP. Accumulation of misfolded proteins in the endoplasmic reticulum causes release of HSPA5/BiP, which allows the luminal domain to homodimerize and promote autophosphorylation of the kinase domain and subsequent activation of its endoribonuclease activity. Its endoribonuclease activity is shown to be specific for XBP1 mRNA and excises 26 nucleotides from XBP1 mRNA. The resulting spliced transcript of XBP1 encodes a transcriptional activator protein that up-regulates expression of UPR target genes. IRE1a is shown to induce transient ER stress signaling and confer a protective effect against pressure overload-induced pathological remodeling in the murine heart. This ZooMAb® recombinant monoclonal antibody, generated by our propriety technology, offers significantly enhanced specificity, affinity, reproducibility, and stability over conventional monoclonals. (Ref.: Steiger, D., et al (2018). J. Biol. Chem. 293(25); 9652-9661). |
| WGK Germany | WGK 1 |
| Storage Temp. | 2-8°C |