ET1705-94

Vinculin Recombinant Rabbit Monoclonal Antibody [JM42-43]

cat.: ET1705-94

Product Type:	Recombinant Rabbit monoclonal IgG, primary antibodies
Species reactivity:	Human, Mouse, Rat, Green monkey, Pig
Applications:	WB, IF-Cell, IF-Tissue, IHC-P
Clonality:	Monoclonal
Clone number:	JM42-43

Form:	Liquid
Storage condition:	Shipped at 4℃. Store at +4℃ short term (1-2 weeks). Store at -20℃ long term.
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: 124 kDa
Isotype:	IgG
Immunogen:	Synthetic peptide within human Vinculin aa 1,000-1,060.
Positive control:	HepG2 cell lysate, HeLa cell lysate, NIH/3T3 cell lysate, C6 cell lysate, Mouse spleen tissue lysate, Rat kidney tissue lysate, NIH/3T3, C6, HeLa, mouse spleen tissue, mouse testis tissue, rat colon tissue, rat spleen tissue, rat testis tissue.
Subcellular location:	Cell junction, Cell membrane, Cytoplasm, Cytoskeleton, Membrane.
Recommended Dilutions: WB IF-Cell IF-Tissue IHC-P	1:20,000-1:50,000 1:1,000 1:500-1:4,000 1:5,000-1:20,000
Uniprot #:	SwissProt: P18206 Human \| Q64727 Mouse \| P85972 Rat
Alternative names:	CMD1W CMH15 Epididymis luminal protein 114 HEL114 Metavinculin MV MVCL OTTHUMP00000019861 OTTHUMP00000019862 VCL VINC VINC_HUMAN Vinculin

Images

Fig1: Western blot analysis of Vinculin on different lysates with Rabbit anti-Vinculin antibody (ET1705-94) at 1/20,000 dilution.

Lane 1: HepG2 cell lysate (15 µg/Lane)
Lane 2: HeLa cell lysate (15 µg/Lane)
Lane 3: NIH/3T3 cell lysate (15 µg/Lane)
Lane 4: C6 cell lysate (15 µg/Lane)
Lane 5: Mouse spleen tissue lysate (20 µg/Lane)
Lane 6: Rat kidney tissue lysate (20 µg/Lane)

Predicted band size: 124 kDa
Observed band size: 124 kDa

Exposure time: 1 minute 21 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 (ET1705-94) at 1/20,000 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: Western blot analysis of Vinculin on different lysates with Rabbit anti-Vinculin antibody (ET1705-94) at 1/20,000 dilution.

Lane 1: Mouse colon tissue lysate
Lane 2: Rat lung tissue lysate
Lane 3: Rat spleen tissue lysate
Lane 4: Rat colon tissue lysate

Lysates/proteins at 10 µg/Lane.

Predicted band size: 124 kDa
Observed band size: 124, ~145 kDa

Exposure time: 2 seconds; ECL: K1801;

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 (ET1705-94) at 1/20,000 dilution was used in K1803 at room temperature for 1 hour. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

Fig3: Western blot analysis of Vinculin on different lysates with Rabbit anti-Vinculin antibody (ET1705-94) at 1/20,000 dilution.

Lane 1: COS-1 (African green monkey kidney fibroblast) cell lysate
Lane 2: Pig brain tissue lysate

Lysates/proteins at 5 µg/Lane.

Predicted band size: 124 kDa
Observed band size: 124 kDa

Exposure time: 17 seconds; ECL: K1801;

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 (ET1705-94) at 1/20,000 dilution was used in K1803 at room temperature for 1 hour. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

Fig4: Western blot analysis of Vinculin on different lysates with Rabbit anti-Vinculin antibody (ET1705-94) at 1/20,000 dilution.

Lane 1: HAP1-parental cell lysate
Lane 2: HAP1-Vinculin KD cell lysate

Lysates/proteins at 10 µg/Lane.

Predicted band size: 124 kDa
Observed band size: 124 kDa

Exposure time: 10 seconds; ECL: K1801;

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 (ET1705-94) at 1/20,000 dilution was used in K1803 at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

	Fig5: Application: Immunocytochemistry (IF-cell) Species: Human Sample: HeLa (Human cervix adenocarcinoma epithelial cell) Fixation: 4% Paraformaldehyde, 15 minutes at room temperature. Permeabilization: 0.1% Triton X-100, 15 minutes at room temperature. Blocking: 1% BSA + 10% normal goat serum, 1 hour at room temperature. Antibody dilution buffer: 1% BSA in PBST. Primary antibody: ET1705-94, 1/1,000, overnight at 4℃. Secondary antibody: Goat Anti-Rabbit IgG (iFluor™ 488, HA1121), 45 minutes at room temperature. Counterstain: Beta tubulin (HA601187, Red), 1/100, overnight at 4℃. The nuclear counterstain was DAPI (Blue).
	Fig6: Immunohistochemical analysis of paraffin-embedded mouse spleen tissue with Rabbit anti-Vinculin antibody (ET1705-94) at 1/20,000 dilution. 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 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (ET1705-94) at 1/20,000 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.
	Fig7: Immunohistochemical analysis of paraffin-embedded mouse testis tissue with Rabbit anti-Vinculin antibody (ET1705-94) at 1/20,000 dilution. 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 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (ET1705-94) at 1/20,000 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 rat colon tissue with Rabbit anti-Vinculin antibody (ET1705-94) at 1/20,000 dilution. 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 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (ET1705-94) at 1/20,000 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 rat spleen tissue with Rabbit anti-Vinculin antibody (ET1705-94) at 1/20,000 dilution. 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 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (ET1705-94) at 1/20,000 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 rat testis tissue with Rabbit anti-Vinculin antibody (ET1705-94) at 1/20,000 dilution. 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 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (ET1705-94) at 1/20,000 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: Flow cytometric analysis of HeLa cells labeling Vinculin.

Cells were fixed and permeabilized. Then stained with the primary antibody (ET1705-94, 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).

Note: All products are “FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE”.