Revision 4

#40435Store at -20C

Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:

WB, IF-IC, FC-FP, ChIP

REACTIVITY:

H M R

SENSITIVITY:

Endogenous

MW (kDa):

60 (human), 55 (mouse/rat)

Source/Isotype:

Rabbit IgG

UniProt ID:

#O35426-2

Entrez-Gene Id:

22433

Product Information

Product Usage Information

For optimal ChIP results, use 10 μl of antibody and 10 μg of chromatin (approximately 4 × 10^6 cells) per IP. This antibody has been validated using SimpleChIP® Enzymatic Chromatin IP Kits.

Application Dilution
Western Blotting 1:1000
Immunofluorescence (Immunocytochemistry) 1:400 - 1:1600
Flow Cytometry (Fixed/Permeabilized) 1:400 - 1:1600
Chromatin IP 1:50

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

For a carrier free (BSA and azide free) version of this product see product #72916.

Specificity / Sensitivity

XBP-1s (E9V3E) Rabbit mAb recognizes endogenous levels of total XBP-1s protein. In western blot analysis, the antibody detects a 78 kDa protein and a 99 kDa protein of unknown identities.

Species Reactivity:

Human, Mouse, Rat

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Ala361 of mouse XBP-1s protein.

Background

Following protein synthesis, secretory, intra-organellar, and transmembrane proteins translocate into the endoplasmic reticulum (ER) where they are post-translationally modified and properly folded. The accumulation of unfolded proteins within the ER triggers an adaptive mechanism known as the unfolded protein response (UPR) that counteracts compromised protein folding (1). The transmembrane serine/threonine kinase IRE1, originally identified in Saccharomyces cerevisiae, is a proximal sensor for the UPR that transmits the unfolded protein signal across the ER membrane (2-4). The human homolog IRE1α was later identified and is ubiquitously expressed in human tissues (5). Upon activation of the unfolded protein response, IRE1α splices X-box binding protein 1 (XBP-1) mRNA through an unconventional mechanism using its endoribonuclease activity (6). This reaction converts XBP-1 from an unspliced XBP-1u isoform to the spliced XBP-1s isoform, which is a potent transcriptional activator that induces expression of many UPR responsive genes (6).

  1. Kaufman, R.J. et al. (2002) Nat Rev Mol Cell Biol 3, 411-21.
  2. Nikawa, J. and Yamashita, S. (1992) Mol Microbiol 6, 1441-6.
  3. Cox, J.S. et al. (1993) Cell 73, 1197-206.
  4. Mori, K. et al. (1993) Cell 74, 743-56.
  5. Tirasophon, W. et al. (1998) Genes Dev 12, 1812-24.
  6. Lee, K. et al. (2002) Genes Dev 16, 452-66.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v nonfat dry milk, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting IF-IC: Immunofluorescence (Immunocytochemistry) FC-FP: Flow Cytometry (Fixed/Permeabilized) ChIP: Chromatin IP

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
Alexa Fluor is a registered trademark of Life Technologies Corporation.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

使用に関する制限

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Revision 4
#40435

XBP-1s (E9V3E) Rabbit mAb

Western Blotting Image 1: XBP-1s (E9V3E) Rabbit mAb Expand Image
Western blot analysis of extracts from 293T cells untreated (-) or treated with Tunicamycin #12819 (2 μg/ml, 8 hr; +), using XBP-1s (E9V3E) Rabbit mAb (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower).
Western Blotting Image 2: XBP-1s (E9V3E) Rabbit mAb Expand Image
Western blot analysis of extracts from C2C12 cells untreated (-) or treated with Tunicamycin #12819 (2 μg/ml, 8 hr; +) and INS-1 cells untreated (-) or treated with Tunicamycin #12819 (5 μg/ml, 24 hr; +), using XBP-1s (E9V3E) Rabbit mAb (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower).
Immunofluorescence Image 1: XBP-1s (E9V3E) Rabbit mAb Expand Image
Confocal immunofluorescent analysis of C2C12 cells, untreated (left) or treated with Tunicamycin #12819 (2 µg/ml, 8 hr; right), using XBP-1s (E9V3E) Rabbit mAb (green). Actin filaments were labeled with DyLight 554 Phalloidin #13054 (red).
Flow Cytometry Image 1: XBP-1s (E9V3E) Rabbit mAb Expand Image
Flow cytometric analysis of C2C12 cells, untreated (blue, low) or treated with tunicamycin (2 μg/mL, 8 hours; green, positive) using XBP-1s (E9V3E) Rabbit mAb (solid lines) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
Chromatin Immunoprecipitation Image 1: XBP-1s (E9V3E) Rabbit mAb Expand Image
Chromatin immunoprecipitations were performed with cross-linked chromatin from C2C12 cells differentiated with 10% horse serum for 7 days then treated with tunicamycin (2 ug/ml) for 7 hours and either XBP-1s (E9V3E) Rabbit mAb #2392 or Normal Rabbit IgG #2729 using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human DNAJB9 Exon 1 Primers #79879, mouse EIF2AK3 promoter primers and SimpleChIP® Mouse GAPDH Intron 2 Primers #8986. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.