EM00704

HSP60 Mouse Monoclonal Antibody [1-80]

cat.: EM00704

Product Type:	Mouse monoclonal IgG1, primary antibodies
Species reactivity:	Human, Mouse, Rat
Applications:	WB, IF-Cell, IHC-P, FC
Clonality:	Monoclonal
Clone number:	1-80

Form:	Liquid
Storage condition:	Shipped at 4℃. Store at +4℃ short term (1-2 weeks). It is recommended to aliquot into single-use upon delivery. Store at -20℃ long term.
Storage buffer:	PBS (pH7.4), 0.1% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.
Concentration:	1ug/ul
Purification:	Protein A affinity purified.
Molecular weight:	Predicted band size: 61 kDa
Isotype:	IgG1
Immunogen:	Synthetic peptide within human HSP60 aa 524-573.
Positive control:	HeLa cell lysate, HepG2 cell lysate, SW480 cell lysate, NIH/3T3 cell lysate, PC-12 cell lysate, Rat kidney tissue lysate, HeLa, NIH/3T3, human kidney tissue, mouse kidney tissue, rat kidney tissue.
Subcellular location:	Mitochondrion matrix.
Recommended Dilutions: WB IF-Cell IHC-P FC	1:5,000 1:1,000 1:1,000 1:1,000
Uniprot #:	SwissProt: P10809 Human \| P63038 Mouse \| P63039 Rat
Alternative names:	60 kDa chaperonin 60 kDa heat shock protein, mitochondrial CH60_HUMAN Chaperonin 60 Chaperonin, 60-KD CPN60 fa04a05 GROEL heat shock 60kDa protein 1 (chaperonin) Heat shock protein 1 (chaperonin) Heat shock protein 60 Heat shock protein 65 heat shock protein family D (Hsp60) member 1 HLD4 Hsp 60 HSP 65 HSP-60 HSP60 HSP65 HSPD1 HuCHA60 Mitochondrial matrix protein P1 P60 lymphocyte protein short heat shock protein 60 Hsp60s1 SPG14

Images

Fig1: Western blot analysis of HSP60 on different lysates with Mouse anti-HSP60 antibody (EM00704) at 1/5,000 dilution.

Lane 1: HeLa cell lysate (20 µg/Lane)
Lane 2: HepG2 cell lysate (20 µg/Lane)
Lane 3: SW480 cell lysate (20 µg/Lane)
Lane 4: NIH/3T3 cell lysate (20 µg/Lane)
Lane 5: PC-12 cell lysate (20 µg/Lane)
Lane 6: Rat kidney tissue lysate (no heat) (40 µg/Lane)

Notice: no heat means the lysate is not boiled.

Predicted band size: 61 kDa
Observed band size: 61 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 (EM00704) at 1/5,000 dilution was used in primary antibody dilution (K1803) at 4℃ overnight. Goat Anti-Mouse IgG - HRP Secondary Antibody (HA1006) at 1/50,000 dilution was used for 1 hour at room temperature.

Fig2: Immunocytochemistry analysis of HeLa cells labeling HSP60 with Mouse anti-HSP60 antibody (EM00704) at 1/1,000 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 Mouse anti-HSP60 antibody (EM00704) at 1/1,000 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Mouse IgG H&L (iFluor™ 488, HA1125) 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 (ET1602-4, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Rabbit IgG H&L (iFluor™ 594, HA1122) were used as the secondary antibody at 1/1,000 dilution.

	Fig3: Immunocytochemistry analysis of NIH/3T3 cells labeling HSP60 with Mouse anti-HSP60 antibody (EM00704) at 1/1,000 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 Mouse anti-HSP60 antibody (EM00704) at 1/1,000 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Mouse IgG H&L (iFluor™ 488, HA1125) 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 (ET1602-4, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Rabbit IgG H&L (iFluor™ 594, HA1122) were used as the secondary antibody at 1/1,000 dilution.
	Fig4: Immunohistochemical analysis of paraffin-embedded human kidney tissue with Mouse anti-HSP60 antibody (EM00704) 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 (EM00704) 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.
	Fig5: Immunohistochemical analysis of paraffin-embedded mouse kidney tissue with Mouse anti-HSP60 antibody (EM00704) 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 (EM00704) 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.

Fig6: Immunohistochemical analysis of paraffin-embedded rat kidney tissue with Mouse anti-HSP60 antibody (EM00704) 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 (EM00704) 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: Flow cytometric analysis of HeLa cells labeling HSP60.

Cells were fixed and permeabilized. Then stained with the primary antibody (EM00704, 1/1,000) (red) compared with Mouse IgG1 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-Mouse IgG Secondary antibody (HA1125) 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”.