WB
Western blot analysis of IKB alpha on different lysates with Rabbit anti-IKB alpha antibody at 1/5,000 dilution. Lane 1: HeLa cell lysate, Lane 2: HeLa cell lysate, Lane 3: SK-Br-3 cell lysate, Lane 4: NIH/3T3 cell lysate, Lane 5: RAW264.7 cell lysate, Lane 6: PC-12 cell lysate, Lysates/proteins at 20 µg/Lane. Exposure time: 1 minute; 12% 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 room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody at 1:300,000 dilution was used for 1 hour at room temperature.IHC
Immunohistochemical analysis of paraffin-embedded mouse stomach tissue using anti-IKB alpha antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (1/50) 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 NIH/3T3 cells labeling IKB alpha with Rabbit anti-IKB alpha 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-IKB alpha antibody at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 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 (iFluor™ 594) was used as the secondary antibody at 1/1,000 dilution.FC
Flow cytometric analysis of HeLa cells labeling IKB alpha. Cells were fixed and permeabilized. Then stained with the primary antibody (1μg/mL) (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).IP
IKB alpha was immunoprecipitated from 0.5 mg Hela whole cell lysates with Rabbit anti-IKB alpha antibody at 2 μg/mL. Western blot was performed from the immunoprecipitate using Rabbit anti-IKB alpha antibody at 1/500 dilution for 45 minutes at room temperature. Goat anti-Rabbit IgG-HRP Secondary Antibody was used at 1:300,000 dilution for 30 minutes at room temperature. Lane 1: Hela whole cell lysates at 10 μg; Lane 2: IKB alpha Rabbit anti-IKB alpha antibody IP in Hela whole cell lysates; Lane 3: Rabbit IgG instead of IKB alpha Rabbit anti-IKB alpha antibody in Hela whole cell lysates. Exposure time: 1 minute; 12% SDS-PAGE gel.| Product Name | IKB alpha Recombinant Rabbit Monoclonal Antibody |
|---|---|
| Antibody Type | Primary Antibodies |
| Product description | NFKB1 or NFKB2 is bound to REL, RELA, or RELB to form the NFKB complex. The NFKB complex is inhibited by I-kappa-B proteins, which inactivate NF-kappa-B by trapping it in the cytoplasm. Phosphorylation of serine residues on the I-kappa-B proteins by kinases marks them for destruction via the ubiquitination pathway, thereby allowing activation of the NF-kappa-B complex. Activated NFKB complex translocates into the nucleus and binds DNA at kappa-B-binding motifs such as 5-prime GGGRNNYYCC 3-prime or 5-prime HGGARNYYCC 3-prime. |
| Immunogen | Synthetic peptide within human IKB alpha aa 1-50. |
| Clonality | monoclonal |
|---|---|
| Isotype | IgG |
| Host Species | Recombinant rabbit |
| Tested Applications | FCICC/IFIHCIPWB |
| WB:1:500-1:5000 IHC:1:50-1:200 IC/IF:1:50-1:200 FC:1:1000 IP:Use at an assay dependent concentration. |
|
| Species Reactivity | HumanMouseRat |
| Concentration | 1mg/ml |
| Purification | Protein A |
| Alternative Names | I kappa B alpha antibody I-kappa-B-alpha antibody IkappaBalpha antibody IkB-alpha antibody IKBA antibody IKBA_HUMAN antibody IKBalpha antibody MAD 3 antibody MAD3 antibody Major histocompatibility complex enhancer-binding protein MAD3 antibody NF kappa B inhibitor alpha antibody NF-kappa-B inhibitor alpha antibody NFKBI antibody NFKBIA antibody Nuclear factor of kappa light chain gene enhancer in B cells antibody Nuclear factor of kappa light polypeptide gene enhancer in B cells inhibitor alpha antibody |
|---|---|
| Molecular Weight(MW) | 36kDa |
| Cellular Localization | Cytoplasm,Nucleus. |
WB
Western blot analysis of IKB alpha on different lysates with Rabbit anti-IKB alpha antibody at 1/5,000 dilution. Lane 1: HeLa cell lysate, Lane 2: HeLa cell lysate, Lane 3: SK-Br-3 cell lysate, Lane 4: NIH/3T3 cell lysate, Lane 5: RAW264.7 cell lysate, Lane 6: PC-12 cell lysate, Lysates/proteins at 20 µg/Lane. Exposure time: 1 minute; 12% 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 room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody at 1:300,000 dilution was used for 1 hour at room temperature.
IHC
Immunohistochemical analysis of paraffin-embedded mouse stomach tissue using anti-IKB alpha antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (1/50) 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 NIH/3T3 cells labeling IKB alpha with Rabbit anti-IKB alpha 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-IKB alpha antibody at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 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 (iFluor™ 594) was used as the secondary antibody at 1/1,000 dilution.
FC
Flow cytometric analysis of HeLa cells labeling IKB alpha. Cells were fixed and permeabilized. Then stained with the primary antibody (1μg/mL) (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).
IP
IKB alpha was immunoprecipitated from 0.5 mg Hela whole cell lysates with Rabbit anti-IKB alpha antibody at 2 μg/mL. Western blot was performed from the immunoprecipitate using Rabbit anti-IKB alpha antibody at 1/500 dilution for 45 minutes at room temperature. Goat anti-Rabbit IgG-HRP Secondary Antibody was used at 1:300,000 dilution for 30 minutes at room temperature. Lane 1: Hela whole cell lysates at 10 μg; Lane 2: IKB alpha Rabbit anti-IKB alpha antibody IP in Hela whole cell lysates; Lane 3: Rabbit IgG instead of IKB alpha Rabbit anti-IKB alpha antibody in Hela whole cell lysates. Exposure time: 1 minute; 12% SDS-PAGE gel.| Positive Control | PC-12, MCF-7, Hela, NIH/3T3, SHG-44, human kidney tissue, mouse brain tissue, mouse kidney tissue, mouse stomach tissue. |
|---|---|
| Application Notes | WB:1:500-1:5000 IHC:1:50-1:200 IC/IF:1:50-1:200 FC:1:1000 IP:Use at an assay dependent concentration. |
| 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. |
