Revision 5

#18640

Store at -20C

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Orders:

877-616-CELL (2355)

[email protected]

Support:

877-678-TECH (8324)

3 Trask Lane | Danvers | Massachusetts | 01923 | USA

For Research Use Only. Not for Use in Diagnostic Procedures.

Applications:
W, IP

Reactivity:
H

Sensitivity:
Endogenous

MW (kDa):
54

Source/Isotype:
Rabbit IgG

UniProt ID:
#Q8NB16

Entrez-Gene Id:
197259

Product Usage Information

Application Dilution
Western Blotting 1:1000
Immunoprecipitation 1:100

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 #42382.

Specificity/Sensitivity

Phospho-MLKL (Ser358) (E7G7P) Rabbit Monoclonal Antibody recognizes endogenous levels of MLKL protein only when phosphorylated at Ser358. This antibody may also bind to MLKL when dually phosphorylated at Thr357 and Ser358.

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Ser358 of human MLKL protein.

Background

Necroptosis, a regulated pathway for necrotic cell death, is triggered by a number of inflammatory signals including cytokines in the tumor necrosis factor (TNF) family, pathogen sensors such as toll-like receptors (TLRs), and ischemic injury (1,2). The process is negatively regulated by caspases and is initiated through a complex containing the RIP1 and RIP3 kinases, typically referred to as the necrosome. Mixed lineage kinase domain-like protein (MLKL) is a pseudokinase that was identified as a downstream target of RIP3 in the necroptosis pathway (3,4). During necroptosis RIP3 is phosphorylated at Ser227, which recruits MLKL and leads to its phosphorylation at Thr357 and Ser358 (3). Knockdown of MLKL through multiple mechanisms results in inhibition of necroptosis (3-5). While the precise mechanism for MLKL-induced necroptosis is unclear, some studies have shown that necroptosis leads to oligomerization of MLKL and translocation to the plasma membrane, where it affects membrane integrity (6-9).

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 BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

W: Western Blotting IP: Immunoprecipitation

Cross-Reactivity Key

H: Human

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.

All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

Limited Uses

Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.

Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 5

Cell Signaling Technology Logo
Western blot analysis of extracts from HT-29 cells, untreated (-) or treated with Z-VAD-FMK (20 μM, added 30 min prior to other compounds; +), human TNF-α (20 ng/mL, 7 hr; +), SM-164 (100 nM, 7 hr; +), or Necrostatin-1s #17802 (1 μM, 7 hr; +), using Phospho-MLKL (Ser358) (E7G7P) Rabbit mAb (upper) or GAPDH (D16H11) XP® Rabbit mAb #5174 (lower).
Western Blotting Image 1: Phospho-MLKL (Ser358) (E7G7P) Rabbit Monoclonal Antibody
Immunoprecipitation of phospho-MLKL (Ser358) protein from HT-29 cell extracts. Cells were treated with Z-VAD-FMK (20 μM, added 30 min prior to other compounds) followed by human TNF-α (20 ng/mL, 7 hr) and SM-164 (100 nM, 7 hr). Lane 1 is 10% input, lane 2 is Rabbit (DA1E) Monoclonal Antibody IgG Isotype Control #3900, and lane 3 is Phospho-MLKL (Ser358) (E7G7P) Rabbit Monoclonal Antibody. Western blot analysis was performed using Phospho-MLKL (Ser358) (E7G7P) Rabbit Monoclonal Antibody. Mouse Anti-rabbit IgG (Conformation Specific) (L27A9) Monoclonal Antibody (HRP Conjugate) #5127 was used as a secondary antibody.
Immunoprecipitation Image 1: Phospho-MLKL (Ser358) (E7G7P) Rabbit Monoclonal Antibody
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.