ET1702-84

NEDD8 Recombinant Rabbit Monoclonal Antibody [JF69-09]

cat.: ET1702-84

Product Type:	Recombinant Rabbit monoclonal IgG, primary antibodies
Species reactivity:	Human, Mouse, Rat, Monkey
Applications:	WB, IF-Cell, IF-Tissue, IHC-P, IP
Clonality:	Monoclonal
Clone number:	JF69-09

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.
Molecular weight:	Predicted band size: 9 kDa
Isotype:	IgG
Immunogen:	Synthetic peptide within Human NEDD8 aa 12-57 / 81.
Positive control:	COS-1 cell lysate, HeLa cell lysate, Raji cell lysate, HepG2 cell lysate, K-562 cell lysate, RAW264.7 cell lysate, NIH/3T3 cell lysate, PC-12 cell lysate, C6 cell lysate, HepG2, human lung carcinoma tissue, human breast carcinoma tissue, human testis tissue, rat bladder tissue, mouse colon tissue, mouse heart tissue, mouse lung tissue, mouse skeletal muscle tissue, mouse spleen tissue, rat brain tissue, rat colon tissue, rat liver tissue, rat lung tissue, rat skeletal muscle tissue, rat testis tissue.
Subcellular location:	Nucleus.
Recommended Dilutions: WB IF-Cell IF-Tissue IHC-P IP	1:500-1:1,000 1:50-1:200 1:50-1:200 1:50-1:1,000 1:50
Uniprot #:	SwissProt: Q15843 Human \| P29595 Mouse \| Q71UE8 Rat
Alternative names:	FLJ43224 MGC104393 MGC125896 MGC125897 NED8 NEDD 8 NEDD-8 Nedd8 NEDD8_HUMAN Neddylin Neural precursor cell expressed developmentally down regulated 8 Neural precursor cell expressed developmentally down regulated gene 8 Neural precursor cell expressed developmentally down-regulated protein 8 Rub1 Ubiquitin like protein Nedd 8 Ubiquitin like protein Nedd8 Ubiquitin-like protein Nedd8

Images

Fig1: Western blot analysis of NEDD8 on different lysates with Rabbit anti-NEDD8 antibody (ET1702-84) at 1/1,000 dilution.

Lane 1: COS-1 cell lysate
Lane 2: HeLa cell lysate
Lane 3: Raji cell lysate
Lane 4: HepG2 cell lysate
Lane 5: K-562 cell lysate
Lane 6: RAW264.7 cell lysate
Lane 7: NIH/3T3 cell lysate
Lane 8: PC-12 cell lysate
Lane 9: C6 cell lysate

Lysates/proteins at 20 µg/Lane.

Predicted band size: 9 kDa
Observed band size: 9-90 kDa

Exposure time: 3 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 (ET1702-84) at 1/1,000 dilution was 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.

Fig2: ICC staining of NEDD8 in HepG2 cells (red). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 10% negative goat serum for 15 minutes at room temperature. Cells were probed with the primary antibody (ET1702-84, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®594 conjugate-Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).

	Fig3: Immunohistochemical analysis of paraffin-embedded human lung carcinoma tissue using anti-NEDD8 antibody. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) 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 (ET1702-84, 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.
	Fig4: Immunohistochemical analysis of paraffin-embedded human breast carcinoma tissue using anti-NEDD8 antibody. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) 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 (ET1702-84, 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.
	Fig5: Immunohistochemical analysis of paraffin-embedded human testis tissue with Rabbit anti-NEDD8 antibody (ET1702-84) at 1/400 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 (ET1702-84) at 1/400 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.
	Fig6: Immunohistochemical analysis of paraffin-embedded rat bladder tissue with Rabbit anti-NEDD8 antibody (ET1702-84) at 1/400 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 (ET1702-84) at 1/400 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.

	Fig7: Immunohistochemical analysis of paraffin-embedded mouse colon tissue with Rabbit anti-NEDD8 antibody (ET1702-84) at 1/1,000 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 (ET1702-84) at 1/1,000 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.
	Fig8: Immunohistochemical analysis of paraffin-embedded mouse heart tissue with Rabbit anti-NEDD8 antibody (ET1702-84) at 1/1,000 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 (ET1702-84) at 1/1,000 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.
	Fig9: Immunohistochemical analysis of paraffin-embedded mouse lung tissue with Rabbit anti-NEDD8 antibody (ET1702-84) at 1/1,000 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 (ET1702-84) at 1/1,000 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.

	Fig10: Immunohistochemical analysis of paraffin-embedded mouse skeletal muscle tissue with Rabbit anti-NEDD8 antibody (ET1702-84) at 1/1,000 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 (ET1702-84) at 1/1,000 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.
	Fig11: Immunohistochemical analysis of paraffin-embedded mouse spleen tissue with Rabbit anti-NEDD8 antibody (ET1702-84) at 1/1,000 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 (ET1702-84) at 1/1,000 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.
	Fig12: Immunohistochemical analysis of paraffin-embedded rat brain tissue with Rabbit anti-NEDD8 antibody (ET1702-84) at 1/1,000 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 (ET1702-84) at 1/1,000 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.

	Fig13: Immunohistochemical analysis of paraffin-embedded rat colon tissue with Rabbit anti-NEDD8 antibody (ET1702-84) at 1/1,000 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 (ET1702-84) at 1/1,000 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.
	Fig14: Immunohistochemical analysis of paraffin-embedded rat liver tissue with Rabbit anti-NEDD8 antibody (ET1702-84) at 1/1,000 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 (ET1702-84) at 1/1,000 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.
	Fig15: Immunohistochemical analysis of paraffin-embedded rat lung tissue with Rabbit anti-NEDD8 antibody (ET1702-84) at 1/1,000 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 (ET1702-84) at 1/1,000 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.

Fig16: Immunohistochemical analysis of paraffin-embedded rat skeletal muscle tissue with Rabbit anti-NEDD8 antibody (ET1702-84) at 1/1,000 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 (ET1702-84) at 1/1,000 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.

Fig17: Immunohistochemical analysis of paraffin-embedded rat testis tissue with Rabbit anti-NEDD8 antibody (ET1702-84) at 1/1,000 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 (ET1702-84) at 1/1,000 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.

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