R1603-5

PKM2 Rabbit Polyclonal Antibody

cat.: R1603-5

Product Type:	Rabbit polyclonal IgG, primary antibodies
Species reactivity:	Human, Mouse, Rat
Applications:	WB, IF-Cell, IHC-P
Clonality:	Polyclonal

Form:	Liquid
Storage condition:	Store at +4℃ after thawing. Aliquot store at -20℃. Avoid repeated freeze / thaw cycles.
Storage buffer:	1*PBS (pH7.4), 0.2% BSA, 50% Glycerol. Preservative: 0.05% Sodium Azide.
Concentration:	1ug/ul
Purification:	Immunogen affinity purified.
Molecular weight:	Predicted band size: 58 kDa
Isotype:	IgG
Immunogen:	Recombinant protein corresponding to C terminal of Human PKM2 .
Positive control:	MDA-MB-231 cell lysate, SiHa cell lysate, NIH/3T3 cell lysate, Rat heart tissue lysate, Rat skeletal muscle tissue lysate, A549, NIH/3T3, PC-12, human lung cancer tissue, human testis tissue, mouse testis tissue, rat testis tissue.
Subcellular location:	Cytoplasm, Nucleus
Recommended Dilutions: WB IF-Cell IHC-P	1:1,000-1:5,000 1:250 1:1,000
Uniprot #:	SwissProt: P14618 Human \| P52480 Mouse \| P11980 Rat
Alternative names:	CTHBP Cytosolic thyroid hormone binding protein Cytosolic thyroid hormone-binding protein KPYM_HUMAN MGC3932 OIP 3 OIP-3 OIP3 OPA interacting protein 3 Opa-interacting protein 3 p58 PK muscle type PK, muscle type PK2 PK3 PKM PKM2 pykm Pyruvate kinase 2/3 Pyruvate kinase 3 Pyruvate kinase isozymes M1/M2 Pyruvate kinase muscle Pyruvate kinase muscle isozyme pyruvate kinase PKM Pyruvate kinase, muscle 2 TCB THBP1 Thyroid hormone binding protein 1 Thyroid hormone binding protein cytosolic Thyroid hormone-binding protein 1 Tumor M2 PK Tumor M2-PK

Images

Fig1: Western blot analysis of PKM2 on different lysates with Rabbit anti-PKM2 antibody (R1603-5) at 1/1,000 dilution.

Lane 1: MDA-MB-231 cell lysate (15 µg/Lane)
Lane 2: SiHa cell lysate (15 µg/Lane)
Lane 3: NIH/3T3 cell lysate (15 µg/Lane)
Lane 4: Rat heart tissue lysate (30 µg/Lane)
Lane 5: Rat skeletal muscle tissue lysate (30 µg/Lane)

Predicted band size: 58 kDa
Observed band size: 58 kDa

Exposure time: 4 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 (R1603-5) 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: All lanes: Western blot analysis of PKM with anti-PKM antibody (R1603-5) at 1:500 dilution.

Lane 1: Wild-type MDA-MB-231 whole cell lysate.
Lane 2: PKM knockout MDA-MB-231 whole cell lysate.

R1603-5 was shown to specifically react with PKM in wild-type MDA-MB-231 cells. No band was observed when PKM knockout sample was tested. Wild-type and PKM knockout samples were subjected to SDS-PAGE. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM in TBST for 1 hour at room temperature. The primary antibody (R1603-5, 1/500) and Loading control antibody (Rabbit anti-GAPDH , ET1601-4, 1/10,000) was used in 5% BSA at room temperature for 2 hours. Goat Anti-Rabbit IgG-HRP Secondary Antibody (HA1001) at 1:200,000 dilution was used for 1 hour at room temperature.

Cell lysate was provided by Ubigene Biosciences (Ubigene Biosciences Co., Ltd., Guangzhou, China).

Fig3: Immunocytochemistry analysis of A549 cells labeling PKM2 with Rabbit anti-PKM2 antibody (R1603-5) at 1/250 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-PKM2 antibody (R1603-5) at 1/250 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.

Fig4: Immunocytochemistry analysis of NIH/3T3 cells labeling PKM2 with Rabbit anti-PKM2 antibody (R1603-5) at 1/250 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-PKM2 antibody (R1603-5) at 1/250 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.

	Fig5: Immunocytochemistry analysis of PC-12 cells labeling PKM2 with Rabbit anti-PKM2 antibody (R1603-5) at 1/250 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-PKM2 antibody (R1603-5) at 1/250 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.
	Fig6: Immunohistochemical analysis of paraffin-embedded human lung cancer tissue with Rabbit anti-PKM2 antibody (R1603-5) 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 (R1603-5) 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.
	Fig7: Immunohistochemical analysis of paraffin-embedded human testis tissue with Rabbit anti-PKM2 antibody (R1603-5) 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 (R1603-5) 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 testis tissue with Rabbit anti-PKM2 antibody (R1603-5) 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 (R1603-5) 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 rat testis tissue with Rabbit anti-PKM2 antibody (R1603-5) 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 (R1603-5) 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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