WB
Western blot analysis of MLKL on different lysates with Rabbit anti-MLKL antibody at 1/5,000 dilution and competitor's antibody at 1/1,000 dilution. Lane 1: HUVEC cell lysate (20 µg/Lane), Lane 2: HT-29 cell lysate (20 µg/Lane), Lane 3: HeLa cell lysate (20 µg/Lane), Lane 4: THP-1 cell lysate (20 µg/Lane), Lane 5: U-937 cell lysate (20 µg/Lane), Lane 6: K-562 cell lysate (low expression) (20 µg/Lane), Lane 7: NIH/3T3 cell lysate (20 µg/Lane), Exposure time: 1 minute 2 seconds; 4-20% SDS-PAGE gel. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody at 1/5,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody at 1/50,000 dilution was used for 1 hour at room temperature.IHC
Immunohistochemical analysis of paraffin-embedded human tonsil tissue using anti-MLKL antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes.The tissues were blocked in 1% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (1/2000) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.ICC/IF
Immunocytochemistry analysis of HeLa cells labeling MLKL with Rabbit anti-MLKL antibody at 1/100 dilution. Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-MLKL antibody at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (488) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI. Beta tubulin (red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (594) was used as the secondary antibody at 1/1,000 dilution.FC
Flow cytometric analysis of HT-29 cells labeling MLKL. Cells were fixed and permeabilized. Then stained with the primary antibody (1/1,000) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).IF-P
Species: Human, Tissue: Tonsil, Sample: Paraffin-embedded section, Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95℃. Wash buffer: 1× PBS, Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes. Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature. Primary antibody: 1/1,000, overnight at 4℃. Secondary antibody: Goat Anti-Rabbit IgG (488), 1.5 hours at room temperature.| Product Name | MLKL Recombinant Rabbit Monoclonal Antibody |
|---|---|
| Antibody Type | Primary Antibodies |
| Product description | MLKL (mixed lineage kinase domain-like) is a 471 amino acid protein that contains one protein kinase domain which is thought to be catalytically inactive. The gene encoding MLKL maps to chromosome 16 and is expressed as two isoforms which are produced by alternative splicing events. Chromosome 16, which is associated with a variety of genetic disorders, encodes over 900 genes and comprises nearly 3% of the human genome. The GAN gene is located on chromosome 16 and, with mutation, may lead to giant axonal neuropathy, a nervous system disorder characterized by increasing malfunction with growth. The rare disorder Rubinstein-Taybi syndrome is associated with chromosome 16, as is Crohn's disease, which is a gastrointestinal inflammatory condition. |
| Immunogen | Recombinant protein within Human MLKL aa 333-471 / 471. |
| Clonality | Monoclonal |
|---|---|
| Isotype | IgG |
| Host Species | Recombinant rabbit |
| Tested Applications | FCICC/IFIF-PIHCWB |
| WB:1:5000-1:10000 IHC:1:500-1:2000 ICC/IF:1:100 FC:1:1000 IF-P:1:500-1:1000 |
|
| Species Reactivity | HumanMouse |
| Concentration | 1mg/ml |
| Purification | Protein A |
| Alternative Names | 9130019I15Rik antibody FLJ34389 antibody hMLKL antibody Mixed lineage kinase domain like antibody Mixed lineage kinase domain like protein antibody Mixed lineage kinase domain-like protein antibody Mlkl antibody MLKL_HUMAN antibody |
|---|---|
| Molecular Weight(MW) | 54kDa |
| Cellular Localization | Cytoplasm, Cell membrane, Golgi apparatus. |
| SwissProt ID | Q8NB16 |
|---|
WB
Western blot analysis of MLKL on different lysates with Rabbit anti-MLKL antibody at 1/5,000 dilution and competitor's antibody at 1/1,000 dilution. Lane 1: HUVEC cell lysate (20 µg/Lane), Lane 2: HT-29 cell lysate (20 µg/Lane), Lane 3: HeLa cell lysate (20 µg/Lane), Lane 4: THP-1 cell lysate (20 µg/Lane), Lane 5: U-937 cell lysate (20 µg/Lane), Lane 6: K-562 cell lysate (low expression) (20 µg/Lane), Lane 7: NIH/3T3 cell lysate (20 µg/Lane), Exposure time: 1 minute 2 seconds; 4-20% SDS-PAGE gel. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody at 1/5,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody at 1/50,000 dilution was used for 1 hour at room temperature.
IHC
Immunohistochemical analysis of paraffin-embedded human tonsil tissue using anti-MLKL antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes.The tissues were blocked in 1% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (1/2000) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
ICC/IF
Immunocytochemistry analysis of HeLa cells labeling MLKL with Rabbit anti-MLKL antibody at 1/100 dilution. Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-MLKL antibody at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (488) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI. Beta tubulin (red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (594) was used as the secondary antibody at 1/1,000 dilution.
FC
Flow cytometric analysis of HT-29 cells labeling MLKL. Cells were fixed and permeabilized. Then stained with the primary antibody (1/1,000) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).
IF-P
Species: Human, Tissue: Tonsil, Sample: Paraffin-embedded section, Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95℃. Wash buffer: 1× PBS, Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes. Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature. Primary antibody: 1/1,000, overnight at 4℃. Secondary antibody: Goat Anti-Rabbit IgG (488), 1.5 hours at room temperature.| Application Notes | WB:1:5000-1:10000 IHC:1:500-1:2000 ICC/IF:1:100 FC:1:1000 IF-P:1:500-1:1000 |
|---|
| Form | Liquid |
|---|---|
| Storage Instructions | Store at +4℃ after thawing. Aliquot store at -20℃ or -80℃. Avoid repeated freeze / thaw cycles. |
| Storage Buffer | 1*TBS (pH7.4), 1%BSA, 40%Glycerol. Preservative: 0.05% Sodium Azide. |
