ET1608-5

mTOR Recombinant Rabbit Monoclonal Antibody [SU30-00]

cat.: ET1608-5

Product Type:	Recombinant Rabbit monoclonal IgG, primary antibodies
Species reactivity:	Human, Mouse, Rat
Applications:	WB, IHC-P, IP, IF-Cell
Clonality:	Monoclonal
Clone number:	SU30-00

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: 289 kDa
Isotype:	IgG
Immunogen:	Synthetic peptide within human mTOR aa 2400-2440.
Positive control:	HeLa cell lysate, MCF7 cell lysate, HEK-293 cell lysate, HepG2 cell lysate, C2C12 cell lysate, PC-12 cell lysate, C6 cell lysate, mouse testis tissue lysate, rat testis tissue lysate, rat heart tissue lysate, SiHa cell lysate, human breast carcinoma tissue, HeLa, NIH/3T3, human kidney tissue, mouse testis tissue, mouse kidney tissue.
Subcellular location:	Endoplasmic reticulum membrane, Microsome membrane, PML body, Golgi apparatus membrane, Cytoplasm, Lysosome, Lysosome membrane, Mitochondrion outer membrane, phagosome.
Recommended Dilutions: WB IHC-P IF-cell IP	1:5,000 1:200-1:1,000 1:50-1:200 Use at an assay dependent concentration.
Uniprot #:	SwissProt: P42345 Human \| Q9JLN9 Mouse \| P42346 Rat
Alternative names:	dJ576K7.1 (FK506 binding protein 12 rapamycin associated protein 1) FK506 binding protein 12 rapamycin associated protein 1 FK506 binding protein 12 rapamycin associated protein 2 FK506 binding protein 12 rapamycin complex associated protein 1 FK506-binding protein 12-rapamycin complex-associated protein 1 FKBP rapamycin associated protein FKBP12 rapamycin complex associated protein FKBP12-rapamycin complex-associated protein 1 FKBP12-rapamycin complex-associated protein FLJ44809 FRAP FRAP1 FRAP2 Mammalian target of rapamycin Mechanistic target of rapamycin mTOR MTOR_HUMAN OTTHUMP00000001983 RAFT1 Rapamycin and FKBP12 target 1 Rapamycin associated protein FRAP2 Rapamycin target protein 1 Rapamycin target protein RAPT1 Serine/threonine-protein kinase mTOR

Images

Fig1: Western blot analysis of mTOR on different lysates with Rabbit anti-mTOR antibody (ET1608-5) at 1/5,000 dilution and competitor's antibody at 1/1,000 dilution.

Lane 1: HeLa cell lysate (20 µg/Lane)
Lane 2: MCF7 cell lysate (20 µg/Lane)
Lane 3: HEK-293 cell lysate (20 µg/Lane)
Lane 4: HepG2 cell lysate (20 µg/Lane)
Lane 5: C2C12 cell lysate (20 µg/Lane)
Lane 6: PC-12 cell lysate (20 µg/Lane)
Lane 7: C6 cell lysate (20 µg/Lane)
Lane 8: Mouse testis tissue lysate (20 µg/Lane)
Lane 9: Rat testis tissue lysate (20 µg/Lane)

Predicted band size: 289 kDa
Observed band size: 289 kDa

Exposure time: 24 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 (ET1608-5) at 1/5,000 dilution and competitor's antibody at 1/1,000 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.

Fig2: Immunocytochemistry analysis of HeLa cells labeling mTOR with Rabbit anti-mTOR antibody (ET1608-5) at 1/100 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-mTOR antibody (ET1608-5) 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 (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.

	Fig3: Immunocytochemistry analysis of NIH/3T3 cells labeling mTOR with Rabbit anti-mTOR antibody (ET1608-5) 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-mTOR antibody (ET1608-5) 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.
	Fig4: Immunohistochemical analysis of paraffin-embedded rat kidney tissue with Rabbit anti-mTOR antibody (ET1608-5) at 1/5,000 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 (ET1608-5) at 1/5,000 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.
	Fig5: Immunohistochemical analysis of paraffin-embedded rat testis tissue with Rabbit anti-mTOR antibody (ET1608-5) at 1/5,000 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 (ET1608-5) at 1/5,000 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.

Fig6: Immunocytochemistry analysis of PC-12 cells labeling mTOR with Rabbit anti-mTOR antibody (ET1608-5) at 1/100 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-mTOR antibody (ET1608-5) 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 (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.

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