Diaminobenzidinef dab

The tumour tissues were xed in 4% paraformaldehyde for 24 hours, followed by dehydrating in a graded alcohol series and embedding in para n, followed by cutting into 5μm sections. The sections were depara nised, rehydrated with a graded alcohol series and then incubated in 96℃ with 0.01 mol/l sodium citrate buffer for the antigen retrieval. Following the incubation in 5% H 2 O 2 for a period of 2 hours, the sections were incubated using primary antibodies including ki67 and SMAD1 (Abcam, England) overnight at 4℃. Immunostaining was carried out with the use of streptavidin-peroxidase and diaminobenzidinef (DAB) following the manufacturer's instructions (Beyotime, Shanghai, China). Eventually, the sections were not only observed under a uorescence microscope (Leica, Wetzlar, Germany) but also imaged.

Tumor tissues were xed in 4% paraformaldehyde for 24 hours, and then dehydrated in alcohol, embedded in para n, and cutted into 5μm sections. The sections were depara nized, rehydrated, and then incubated at 96℃ with 0.01 mol/l sodium citrate buffer for the antigen retrieval. Following the incubation in 5% H 2 O 2 for 2 hours, the sections were incubated with primary antibodies overnight at 4℃.
Immunostaining was carried out with streptavidin-peroxidase and diaminobenzidinef (DAB) following the manufacturer's instructions (Beyotime, Shanghai, China). Finally, the sections were observed and imaged under uorescence microscope (Leica, Wetzlar, Germany).