Ubiquitin Recombinant Rabbit Monoclonal Antibody [ST46-03]
cat.: ET1609-21
Product Type: Recombinant Rabbit monoclonal IgG, primary antibodies
Species reactivity: Human, Mouse, Rat
Applications: WB, IF-Cell, IF-Tissue, IHC-P, FC
Clonality: Monoclonal
Clone number: ST46-03
Form: Liquid
Storage condition: Store at +4℃ after thawing. Aliquot store at -20℃ or -80℃. Avoid repeated freeze / thaw cycles.
Storage buffer: 1*TBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.
Concentration: 1ug/ul
Purification: Protein A affinity purified.
Isotype: IgG
Immunogen: Synthetic peptide within human Ubiquitin aa 1-50.
Positive control: HeLa cell lysate, HeLa treated with 10μM MG-132 for 6 hours cell lysate, NIH/3T3 cell lysate, NIH/3T3 treated with 10μM MG-132 for 8 hours cell lysate, PC-12 cell lysate, HepG2, mouse testis tissue, mouse placenta tissue, mouse brain tissue, human esophagus tissue, 293T cell lysate, HepG2 cell lysate.
Subcellular location: Cytoplasm, Nucleus, Membrane, Mitochondrion.
Recommended Dilutions:
  WB
  IF-Cell
  IF-Tissue
  IHC-P
  FC

1:1,000-1:5,000
1:200-1:500
1:50-1:2,000
1:200-1:1,0000
1:1,000
Uniprot #: SwissProt: P0CG47 Human | P0CG48 Human | P62979 Human | P62987 Human | P0CG49 Mouse | P62991 Mouse | P0CG51 Rat | Q63429 Rat
Alternative names: FLJ25987 MGC8385 Polyubiquitin B RPS 27A RPS27A UBA 52 UBA 80 UBA52 UBA80 UBB UBB_HUMAN UBC UBCEP 1 UBCEP 2 UBCEP1 UBCEP2 Ubiquitin Ubiquitin B
Images
ET1609-21_1.jpg Fig1: Western blot analysis of Ubiquitin on different lysates with Rabbit anti-Ubiquitin antibody (ET1609-21) at 1/5,000 dilution and competitor's antibody at 1/2,000 dilution.

Lane 1: HeLa cell lysate
Lane 2: HeLa treated with 10μM MG-132 for 6 hours cell lysate
Lane 3: NIH/3T3 cell lysate
Lane 4: NIH/3T3 treated with 10μM MG-132 for 8 hours cell lysate
Lane 5: PC-12 cell lysate

Lysates/proteins at 20 µg/Lane.

Exposure time: 1 minute 34 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 (ET1609-21) at 1/5,000 dilution and competitor's antibody at 1/2,000 dilution were used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
ET1609-21_2.jpg Fig2: Immunocytochemistry analysis of HepG2 cells labeling Ubiquitin with Rabbit anti-Ubiquitin antibody (ET1609-21) at 1/500 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-Ubiquitin antibody (ET1609-21) at 1/500 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) 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 (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.
ET1609-21_3.png Fig3: Western blot analysis of Ubiquitin on different lysates with Rabbit anti-Ubiquitin antibody (ET1609-21) at 1/1,000 dilution.

Lane 1: PC-12 cell lysate
Lane 2: HeLa cell lysate
Lane 3: NIH/3T3 cell lysate
Lane 4: 293T cell lysate
Lane 5: HepG2 cell lysate

Lysates/proteins at 20 µg/Lane.

Exposure time: 2 minutes;

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 (ET1609-21) at 1/1,000 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/100,000 dilution was used for 1 hour at room temperature
ET1609-21_4.jpg Fig4: Flow cytometric analysis of HepG2 cells labeling Ubiquitin.

Cells were fixed and permeabilized. Then stained with the primary antibody (ET1609-21, 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 (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).
ET1609-21_5.jpg Fig5: Immunohistochemical analysis of paraffin-embedded human esophagus tissue with Rabbit anti-Ubiquitin antibody (ET1609-21) at 1/10,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1609-21) at 1/10,000 dilution for 1 hour 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.
ET1609-21_6.jpg Fig6: Immunohistochemical analysis of paraffin-embedded mouse esophagus tissue with Rabbit anti-Ubiquitin antibody (ET1609-21) at 1/10,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1609-21) at 1/10,000 dilution for 1 hour 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.
ET1609-21_7.jpg Fig7: Immunohistochemical analysis of paraffin-embedded rat esophagus tissue with Rabbit anti-Ubiquitin antibody (ET1609-21) at 1/10,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1609-21) at 1/10,000 dilution for 1 hour 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.
ET1609-21_8.jpg Fig8: Immunohistochemical analysis of paraffin-embedded mouse brain tissue with Rabbit anti-Ubiquitin antibody (ET1609-21) at 1/10,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1609-21) at 1/10,000 dilution for 1 hour 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.
ET1609-21_9.jpg Fig9: Immunohistochemical analysis of paraffin-embedded rat brain tissue with Rabbit anti-Ubiquitin antibody (ET1609-21) at 1/10,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1609-21) at 1/10,000 dilution for 1 hour 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.
ET1609-21_10.jpg Fig10: Immunocytochemistry analysis of NIH/3T3 cells labeling Ubiquitin with Rabbit anti-Ubiquitin antibody (ET1609-21) at 1/500 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-Ubiquitin antibody (ET1609-21) at 1/500 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) 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 (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.
ET1609-21_11.jpg Fig11: Immunocytochemistry analysis of PC-12 cells labeling Ubiquitin with Rabbit anti-Ubiquitin antibody (ET1609-21) at 1/200 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-Ubiquitin antibody (ET1609-21) at 1/200 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) 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 (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.
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