Anti-Hsc70 Antibody Picoband®(monoclonal, 3B6)

Western blot analysis of HSC70/HSPA8 using anti-HSC70/HSPA8 antibody (M01024-1).
Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 30 ug of sample under reducing conditions.
Lane 1: human Caco-2 whole cell lysates,
Lane 2: human A549 whole cell lysates,
Lane 3: human PC-3 whole cell lysates,
Lane 4: human Hacat whole cell lysates,
Lane 5: human U251 whole cell lysates,
Lane 6: human THP-1 whole cell lysates,
Lane 7: human SiHa whole cell lysates,
Lane 8: human MCF-7 whole cell lysates.
After electrophoresis, proteins were transferred to a membrane. Then the membrane was incubated with mouse anti-HSC70/HSPA8 antigen affinity purified monoclonal antibody (M01024-1) at 1:500 overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-mouse IgG-HRP secondary antibody at a dilution of 1:500 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1002) with Tanon 5200 system. A specific band was detected for HSC70/HSPA8 at approximately 71 kDa. The expected band size for HSC70/HSPA8 is at 71 kDa.

Western blot analysis of HSC70/HSPA8 using anti-HSC70/HSPA8 antibody (M01024-1).
Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 30 ug of sample under reducing conditions.
Lane 1: rat thymus tissue lysates,
Lane 2: rat stomach tissue lysates,
Lane 3: rat brain tissue lysates,
Lane 4: rat RH-35 whole cell lysates,
Lane 5: mouse thymus tissue lysates,
Lane 6: mouse stomach tissue lysates,
Lane 7: mouse brain tissue lysates,
Lane 8: mouse NIH/3T3 whole cell lysates.
After electrophoresis, proteins were transferred to a membrane. Then the membrane was incubated with mouse anti-HSC70/HSPA8 antigen affinity purified monoclonal antibody (M01024-1) at 1:500 overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-mouse IgG-HRP secondary antibody at a dilution of 1:500 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1002) with Tanon 5200 system. A specific band was detected for HSC70/HSPA8 at approximately 71 kDa. The expected band size for HSC70/HSPA8 is at 71 kDa.

IHC analysis of Hsc70 using anti-Hsc70 antibody (M01024-1).
Hsc70 was detected in paraffin-embedded section of human lung cancer tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml mouse anti-Hsc70 Antibody (M01024-1) overnight at 4°C. Biotinylated goat anti-mouse IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1021) with DAB as the chromogen.

IHC analysis of Hsc70 using anti-Hsc70 antibody (M01024-1).
Hsc70 was detected in paraffin-embedded section of human rectal cancer (lymph node) tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml mouse anti-Hsc70 Antibody (M01024-1) overnight at 4°C. Biotinylated goat anti-mouse IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1021) with DAB as the chromogen.

IHC analysis of Hsc70 using anti-Hsc70 antibody (M01024-1).
Hsc70 was detected in paraffin-embedded section of human rectal cancer (lymph node) tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml mouse anti-Hsc70 Antibody (M01024-1) overnight at 4°C. Biotinylated goat anti-mouse IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1021) with DAB as the chromogen.

IHC analysis of Hsc70 using anti-Hsc70 antibody (M01024-1).
Hsc70 was detected in paraffin-embedded section of human tonsil tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml mouse anti-Hsc70 Antibody (M01024-1) overnight at 4°C. Biotinylated goat anti-mouse IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1021) with DAB as the chromogen.

IHC analysis of Hsc70 using anti-Hsc70 antibody (M01024-1).
Hsc70 was detected in paraffin-embedded section of rat spleen tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml mouse anti-Hsc70 Antibody (M01024-1) overnight at 4°C. Biotinylated goat anti-mouse IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1021) with DAB as the chromogen.

IHC analysis of Hsc70 using anti-Hsc70 antibody (M01024-1).
Hsc70 was detected in a paraffin-embedded section of human liver cancer tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1:200 mouse anti-Hsc70 Antibody (M01024-1) overnight at 4°C. Peroxidase Conjugated Goat Anti-mouse IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using HRP Conjugated Mouse IgG Super Vision Assay Kit (Catalog # SV0001) with DAB as the chromogen.

IF analysis of Hsc70 using anti-Hsc70 antibody (M01024-1).
Hsc70 was detected in immunocytochemical section of A549 cells. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent (AR0022) for 15 mins. The cells were blocked with 10% goat serum. And then incubated with 2μg/mL mouse anti-Hsc70 Antibody (M01024-1) overnight at 4°C. DyLight®594 Conjugated Goat Anti-Mouse IgG (BA1141) was used as secondary antibody at 1:100 dilution and incubated for 30 minutes at 37°C. The section was counterstained with DAPI. Visualize using a fluorescence microscope and filter sets appropriate for the label used.

Flow Cytometry analysis of A549 cells using anti- Hsc70 antibody (M01024-1).
Overlay histogram showing A549 cells stained with M01024-1 (Blue line). To facilitate intracellular staining, cells were fixed with 4% paraformaldehyde and permeabilized with permeabilization buffer. The cells were blocked with 10% normal goat serum. And then incubated with mouse anti- Hsc70 Antibody (M01024-1, 1μg/1x10⁶ cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-mouse IgG (BA1126, 5-10μg/1x10⁶ cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was mouse IgG (1μg/1x10⁶) used under the same conditions. Unlabelled sample without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.

Flow Cytometry analysis of K562 cells using anti- Hsc70 antibody (M01024-1).
Overlay histogram showing K562 cells stained with M01024-1 (Blue line). To facilitate intracellular staining, cells were fixed with 4% paraformaldehyde and permeabilized with permeabilization buffer. The cells were blocked with 10% normal goat serum. And then incubated with mouse anti- Hsc70 Antibody (M01024-1, 1μg/1x10⁶ cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-mouse IgG (BA1126, 5-10μg/1x10⁶ cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was mouse IgG (1μg/1x10⁶) used under the same conditions. Unlabelled sample without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.

Flow Cytometry analysis of HEPA1-6 cells using anti- Hsc70 antibody (M01024-1).
Overlay histogram showing HEPA1-6 cells stained with M01024-1 (Blue line). To facilitate intracellular staining, cells were fixed with 4% paraformaldehyde and permeabilized with permeabilization buffer. The cells were blocked with 10% normal goat serum. And then incubated with mouse anti- Hsc70 Antibody (M01024-1, 1μg/1x10⁶ cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-mouse IgG (BA1126, 5-10μg/1x10⁶ cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was mouse IgG (1μg/1x10⁶) used under the same conditions. Unlabelled sample without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.