Antibodies, Proteins, Kits and Reagents for Life Science

348 441 ₸ Product size

100 µl 348 441 ₸

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Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091)

Key features and details

Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
Rabbit monoclonal [EPR9514] to MLKL (phospho S358)
Suitable for: WB, Dot blot, IHC-P
Reacts with: Human

Product name

Anti-MLKL (phospho S358) antibody [EPR9514]
See all MLKL primary antibodies
Description

Rabbit monoclonal [EPR9514] to MLKL (phospho S358)
Host species

Rabbit
Specificity

Stimulation may be required to allow detection of the phosphorylated protein. Please see images below for recommended treatment conditions and positive controls.
Tested Applications & Species

Application Species
IHC-P
Human

WB
Human

See all applications and species data
Immunogen

This information is proprietary to Abcam and/or its suppliers.
(Peptide available as ab206929)
Positive control
- WB: HT-29 cell lysate treated with TNF alpha+ Smac mimetic+ z-VAD. HT-29 cells were treated with the indicated stimuli for 8 hours and then harvested. The final concentrations of 20 ng/ml TNF-a, 100 nM Smac mimetic, and 20 µM z-VAD were used to induce necrosis; human hepatocyte (treated with Smac/z0VAD) cell lysate. IHC-P: Human skin and melanoma tissue. Dot blot: MLKL (pS358) phospho peptide; MLKL (pT357/pS358) phospho peptide.
General notes
This antibody was developed through collaboration with the lab of Xiaodong Wang at the National Institute of Biological Sciences, Beijing.
Abcam recommended secondaries - Goat Anti-Rabbit HRP (ab205718) and Goat Anti-Rabbit Alexa Fluor^® 488 (ab150077).
See other anti-rabbit secondary antibodies that can be used with this antibody.
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
For more information see here.
Our RabMAb^® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb^® patents.

Application	Species
IHC-P	Human
WB	Human

Form

Liquid
Storage instructions

Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.
Storage buffer

pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol, 0.05% BSA
Concentration information loading...
Purity

Protein A purified
Clonality

Monoclonal
Clone number

EPR9514
Isotype

IgG
Research areas

Western blot - Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091)

All lanes : Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091) at 1/1000 dilution

Lane 1 : Untreated HT-29 (human colorectal adenocarcinoma) whole cell lysates 20µg
Lane 2 : HT-29 (human colorectal adenocarcinoma) treated with TNF alpha+ Smac mimetic+ z-VAD whole cell lysates 20µg
Lane 3 : HT-29 (human colorectal adenocarcinoma) treated with TNF alpha+ Smac mimetic + z-VAD and phosphatase whole cell lysates 20µg.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

Predicted band size: 54 kDa
Observed band size: 54 kDa

Blocking buffer and concentration: 5% NFDM/TBST, Diluting buffer and concentration: 5% NFDM /TBST, Exposure time: 1 minute

The lysate in this image is prepared by 1%SDS Hot Lysate buffer.

For Lysate preparation protocol, please refer to the protocol book in the protocol section and/or here (downloadable copy).
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091)

ab187091 at 1:250 staining MLKL (phospho S358) in Human melanoma tissue by immunohistochemistry (FFPE).

Antigen retrival required on FFPE tissue: HIER using 10mM Citrate buffer pH 6.0, see recommended HIER protocol

For additional IHC guideline, please see IHC resource page
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091)

ab187091 at 1:250 staining MLKL (phospho S358) in Human skin tissue by immunohistochemistry (FFPE).

Antigen retrival required on FFPE tissue: HIER using 10mM Citrate buffer pH 6.0, see recommended HIER protocol
For additional IHC guideline, please see IHC resource page
Dot Blot - Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091)

Dot blot analysis of MLKL peptides using ab187091 at 1/1000 dilution followed by Goat Anti-Rabbit IgG, (H+L),Peroxidase conjugated secondary antibody at 1/1000 dilution. Blocking and diluting buffer was 5% NFDM/TBST, exposure time 3 minuts.

Lane 1: MLKL (pT357) phospho peptide

Lane 2: MLKL (pS358) phospho peptide

Lane 3: MLKL (pT357/pS358) phospho peptide

Lane 4: MLKL non-phospho peptide
Western blot - Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091) Data courtesy of an anonymous AbReview

All lanes : Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091) at 1000 cells

Lane 1 : Untreated human hepatocyte cell lysate
Lane 2 : Treated (10 ng/ml TNF-a+100 nM Smac mimetic+20 µM z-VAD 6 h) human hepatocyte cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat anti-Rabbit HRP at 1/5000 dilution

Predicted band size: 54 kDa

See Abreview
Western blot - Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091)

Sample: HT-29 (Human colorectal adenocarcinoma epithelial cell) treated with 20 ng/ml TNF-a, 100 nM Smac mimetic and 20 µM z-VAD for 8 hours whole cell lysates 10 µg per lane.

Lane 1 : Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091) at 0.12 µg/ml (Batch produced in 2016)
Lane 2 : Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091) at 0.17 µg/ml (Batch produced in 2015)
Lane 3 : Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091) at 0.12 µg/ml (GR212667 - batch produced in 2014)
Lane 4 : Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091) at 0.16 µg/ml (The supernatant of the clone producing ab187091)
Lane 5 : Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091) at 0.15 µg/ml (Batch produced in 2017)

Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

Blocking and diluting buffer: 5% NFDM/TBST.

The lysate in this image is prepared by 1%SDS Hot Lysate buffer.

For Lysate preparation protocol, please refer to the protocol book in the protocol section and/or here (downloadable copy).
Western blot - Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091)

All lanes : Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091) at 1/5000 dilution

Lane 1 : HT-29 (Human colorectal adenocarcinoma epithelial cell) treated with 20 ng/ml TNF-a, 100 nM Smac mimetic and 20 µM z-VAD for 6 hr. The lysate is directly prepared by 1xSDS loading buffer.
Lane 2 : HT-29 (Human colorectal adenocarcinoma epithelial cell) treated with 20 ng/ml TNF-a, 100 nM Smac mimetic and 20 µM z-VAD for 8 hr. The lysate is prepared by 1%SDS Hot Lysate buffer method.

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

Predicted band size: 54 kDa
Observed band size: 54 kDa

Exposure time: 3 minutes

Blocking and diluting buffer: 5% NFDM/TBST.

For 1%SDS Hot Lysate preparation protocol, please refer to the protocol book in the protocol section and/or here (downloadable copy).
Western blot - Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091)

All lanes : Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091) at 1/2000 dilution

Lane 1 : Untreated HT-29 lysate
Lane 2 : HT-29 cell lysate treated with TNF alpha+ Smac mimetic+ z-VAD

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

Predicted band size: 54 kDa

Details on WB tested positive control samples: HT-29 cells were treated with the indicated stimuli for 8 hr and then harvested. The final concentrations of 20 ng/ml TNF-a, 100 nM Smac mimetic, and 20 µM z-VAD were used to induce necrosis.

The lysate in this image is prepared by 1%SDS Hot Lysate buffer.

For Lysate preparation protocol, please refer to the protocol book in the protocol section and/or here (downloadable copy).
Anti-MLKL (phospho S358) antibody [EPR9514] (ab187091)

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