Super sensitive horseradish peroxidase immunohistochemistry kit

Cells and tissues were fixed with 4% paraformaldehyde for 30 min, washed with PBS three times, permeabilized and blocked with 10% normal goat serum containing 0.3% Triton X-100 and 1% BSA for 2 h, and then incubated with primary antibody overnight at 4 °C. For immunofluorescence, cells and tissues were washed three times with PBS and incubated with the corresponding fluorescent secondary antibody at room temperature for 2 h. Nuclei were counterstained with Hoechst 33342 (1:1000; Pierce, Rockford, IL, USA). Primary antibodies included anti-Tuj1 (1:1000; Millipore) and anti-MAP2 (1:1000; Abcam). Images were captured by using a fluorescence microscope.
Immunohistochemistry was performed using a Super-Sensitive Horseradish Peroxidase Immunohistochemistry Kit (rabbit; Sangon Biotech, Shanghai, China). Sections were incubated with rabbit anti-Ndel1 antibody (1:1000, Abcam) at 4 °C overnight followed by incubation with poly-HRP-conjugated anti-rabbit IgG. After rinsing in PBS, sections were detected using a DAB working solution.

Paraffin sections (5 μm) from tumor tissue samples were deparaffinized in 100% xylene and rehydrated with a decreasing ethanol series and then water. Antigen retrieval was conducted in 0.01 mol/L sodium citrate buffer (pH 6.0) at 100 °C for 10 min. Immunohistochemistry assay was performed as described previously [22 (link)] using super-sensitive horseradish peroxidase immunohistochemistry kit (Sangon Biotech, Shanghai, China) per the manufacturer’s instructions, and the anti-Ki67 antibody (sc-23900, Santa Cruz) was used.

Immunohistochemistry analysis was performed as described previously^{40 (link)}, using a super-sensitive horseradish peroxidase immunohistochemistry kit (Sangon Biotech, Shanghai, China); the anti-Ki67 (sc-23900, Santa Cruz), anti-IGF1R (D163034, BBI), and anti-IRS1 (D120888, BBI) antibodies were used.