ET1702-75

PDHA1 Recombinant Rabbit Monoclonal Antibody [JF996-0]

cat.: ET1702-75

Product Type:	Recombinant Rabbit monoclonal IgG, primary antibodies
Species reactivity:	Human, Mouse, Rat
Applications:	WB, IF-Cell, IF-Tissue, IHC-P, IP, FC
Clonality:	Monoclonal
Clone number:	JF996-0

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: 43 kDa
Isotype:	IgG
Immunogen:	Synthetic peptide within Human PDHA1 aa 100-145 / 390.
Positive control:	A431 cell lysate, C2C12 cell lysate, L6 cell lysate, Mouse heart tissue lysate, Rat heart tissue lysate, A431, C2C12, human kidney tissue, mouse kidney tissue, rat kidney tissue, mouse skeletal muscle tissue.
Subcellular location:	Mitochondrion matrix.
Recommended Dilutions: WB IF-Cell IF-Tissue IHC-P FC IP	1:2,000 1:100-1:500 1:100-1:500 1:200-1:1,000 1:1,000 Use at an assay dependent concentration.
Uniprot #:	SwissProt: P08559 Human \| P35486 Mouse \| P26284 Rat
Alternative names:	ODPA_HUMAN PDH PDHA PDHA1 PDHCE1A PDHE1 A type I PDHE1-A type I PHE1A Pyruvate Dehydrogenase (lipoamide) alpha 1 Pyruvate dehydrogenase complex, E1 alpha polypeptide 1 Pyruvate Dehydrogenase E1 alpha Pyruvate dehydrogenase E1 component subunit alpha, somatic form, mitochondrial

Images

Fig1: Western blot analysis of PDHA1 on different lysates with Rabbit anti-PDHA1 antibody (ET1702-75) at 1/2,000 dilution.

Lane 1: A431 cell lysate (20 µg/Lane)
Lane 2: C2C12 cell lysate (20 µg/Lane)
Lane 3: L6 cell lysate (20 µg/Lane)
Lane 4: Mouse heart tissue lysate (40 µg/Lane)
Lane 5: Rat heart tissue lysate (40 µg/Lane)

Predicted band size: 43 kDa
Observed band size: 43 kDa

Exposure time: Lane 1-3: 30 seconds; Lane 4-5: 12 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 (ET1702-75) at 1/2,000 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 PDHA1 on different lysates with Rabbit anti-PDHA1 antibody (ET1702-75) at 1/1,000 dilution.

Lane 1: HAP1-parental cell lysate
Lane 2: HAP1-PDHA1 KD cell lysate

Lysates/proteins at 10 µg/Lane.

Predicted band size: 43 kDa
Observed band size: 43 kDa

Exposure time: 40 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 (ET1702-75) at 1/1,000 dilution was used in primary antibody dilution (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.

Fig3: Immunocytochemistry analysis of A431 cells labeling PDHA1 with Rabbit anti-PDHA1 antibody (ET1702-75) at 1/500 dilution.

Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 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-PDHA1 antibody (ET1702-75) 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 (HA601187, 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.

Fig4: Immunocytochemistry analysis of C2C12 cells labeling PDHA1 with Rabbit anti-PDHA1 antibody (ET1702-75) at 1/100 dilution.

Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 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-PDHA1 antibody (ET1702-75) at 1/100 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 (HA601187, 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.

	Fig5: Immunohistochemical analysis of paraffin-embedded human kidney tissue with Rabbit anti-PDHA1 antibody (ET1702-75) at 1/200 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 (ET1702-75) at 1/200 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 mouse kidney tissue with Rabbit anti-PDHA1 antibody (ET1702-75) at 1/200 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 (ET1702-75) at 1/200 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 rat kidney tissue with Rabbit anti-PDHA1 antibody (ET1702-75) at 1/200 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 (ET1702-75) at 1/200 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 skeletal muscle tissue with Rabbit anti-PDHA1 antibody (ET1702-75) at 1/1,000 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 (ET1702-75) 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: Flow cytometric analysis of A431 cells labeling PDHA1.

Cells were fixed and permeabilized. Then stained with the primary antibody (ET1702-75, 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 (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”.