Merck, 17-10400, Anti-phospho-MLKL (Ser358) Antibody Set

General description
Mixed lineage kinase domain-like protein (MLKL) is required for the execution of programmed necrosis. MLKL is a cell-permeable acrylamide compound that inhibits human, but not murine, MLKL adaptor function via covalent modification of Cys86. MLKL prevents the MLKL-RIP1-RIP3 necrosome complex from interacting with further downstream effectors.
This phospho-MLKL (Ser358) antibody set contains one vial of phospho-MLKL (Ser358) antibody and one vial of MLKL “Total” antibody.

RECOMMENDED USAGE:
It is recommended to first perform immunoprecipitation on your sample using the non-phosphorylated MLKL antibody for “total” MLKL protein isolation (see protocol section on page 2 of the Certificate of Analysis); then use the immunoprecipitated sample to detect the phosphorylated protein using the phospho-MLKL (Ser358) antibody via western blot.
~54 kDa
Immunogen
Epitope: Ser358
Linear peptide corresponding to the Protein Kinase Domain of phospho-MLKL (Ser358).
Application
Western Blot Analysis:
Untreated HT-29 cell lysates or lysates from HT-29 cells treated with Cycloheximide,Z-VAD-FMK and TNF alpha were first immunoprecipitated using the Anti-MLKL “total” antibody (1:1000 dilution) to isolate the MLKL protein.
The isolated “total” MLKL protein sample was then probed with Anti-phospho-MLKL (Ser358) (1:1000 dilution).
Anti-phospho-MLKL (Ser358) Antibody Set is an antibody set against phospho-MLKL (Ser358) for use in WB.
Biochem/physiol Actions
When used together and under the “Recommended Usage” instructions, this antibody set will detect MLKL phosphorylated at Ser358.
Physical form
100 µL of Anti-phospho-MLKL (Ser358) purified polyclonal antibody in 0.1 M Tris-Glycine (pH 7.4), 150 mM NaCl, with 0.05% sodium azide.

100 µg (concentration: 1 mg/mL) of Anti- MLKL purified polyclonal antibody in 0.1 M Tris-Glycine (pH 7.4), 150 mM NaCl, with 0.05% sodium azide.
Analysis Note
Anti- MLKL (Total Antibody): Evaluated by Immunoprecipitation on HT-29 cell lysates.

Anti-phospho-MLKL (Ser358) Antibody: Evaluated by Dot Blot on phosphorylated and non-phosphorylated MLKL peptides.
Other Notes
Concentration: Please refer to the Certificate of Analysis for the lot-specific concentration.
One vial of phospho-MLKL (Ser358) antibody

One vial of MLKL “Total” antibody