Anti-Hsp70 antibody [2A4] (ab5442)

IHC image of Hsp70 staining in human lung formalin fixed paraffin embedded tissue section*. The section was pre-treated using pressure cooker heat mediated antigen retrieval with sodium citrate buffer (pH6) for 30mins. The section was incubated with ab5442, 1/2000 dilution overnight at +4°C. An HRP-conjugated secondary (ab97230, 1/2000 dilution) was used for 1hr at room temperature. The section was counterstained with haematoxylin and mounted with DPX.

The inset negative control image is secondary-only at 1/500 dilution.

*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

Immunocytochemistry/Immunofluorescence analysis of Hsp70 (green) in Hela cells. Formalin-fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 5-10 minutes at room temperature and blocked with 3% BSA-PBS for 30 minutes at room temperature. Cells were probed with ab5442 at a dilution of 1:100 and incubated overnight in a humidified chamber. Cells were washed with PBST and incubated with a DyLight-conjugated secondary antibody for 45 minutes at room temperature in the dark. F-actin (red) was stained with a fluorescent phalloidin and nuclei (blue) were stained with DAPI. Images were taken at a 60X magnification.

Immunocytochemistry/Immunofluorescence analysis of Hsp70 (green) in A431 cells. Formalin-fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 5-10 minutes at room temperature and blocked with 3% BSA-PBS for 30 minutes at room temperature. Cells were probed with ab5442 at a dilution of 1:100 and incubated overnight in a humidified chamber. Cells were washed with PBST and incubated with a DyLight-conjugated secondary antibody for 45 minutes at room temperature in the dark. F-actin (red) was stained with a fluorescent phalloidin and nuclei (blue) were stained with DAPI. Images were taken at a 60X magnification.

Immunocytochemistry/Immunofluorescence analysis of Hsp70 (green) in NIH-3T3 cells. Formalin-fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 5-10 minutes at room temperature and blocked with 3% BSA-PBS for 30 minutes at room temperature. Cells were probed with ab5442 at a dilution of 1:200 and incubated overnight in a humidified chamber. Cells were washed with PBST and incubated with a DyLight-conjugated secondary antibody for 45 minutes at room temperature in the dark. F-actin (red) was stained with a fluorescent phalloidin and nuclei (blue) were stained with DAPI. Images were taken at a 60X magnification.

Immunocytochemistry/Immunofluorescence analysis of Hsp70 in HeLa Cells. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were incubated without (control) or with ab5442 at a dilution of 1:200 overnight at 4 C, washed with PBS and incubated with a DyLight-488 conjugated secondary antibody. Hsp70 staining (green), F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Images were taken at 60X magnification.

Immunocytochemistry/Immunofluorescence analysis of Hsp70 in NCI-H1299 Cells. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control) or with ab5442 at a dilution of 1:100 overnight at 4 C, washed with PBS and incubated with a DyLight-488 conjugated secondary antibody. Hsp70 staining (green), F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Images were taken at 60X magnification.

Immunocytochemistry/Immunofluorescence analysis of Hsp70 in NIH-3T3 Cells. Cells were grown on chamber slides and fixed with formaldehyde prior to staining. Cells were probed without (control) or with ab5442 at a dilution of 1:100 overnight at 4 C, washed with PBS and incubated with a DyLight-488 conjugated secondary antibody. Hsp70 staining (green), F-Actin staining with Phalloidin (red) and nuclei with DAPI (blue) is shown. Images were taken at 60X magnification.

Immunocytochemistry/Immunofluorescence analysis of Hsp70 (green) in HeLa and NIH3T3 cells. Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with ab5442 at a dilution of 1:50 for at least 1 hour at room temperature, washed with PBS, and incubated with fluorescently labeled goat anti-mouse IgM secondary antibody at a dilution of 1:400 for 30 minutes at room temperature. Nuclei (blue) were stained with Hoechst 33342 dye. Images were taken at 20X magnification.