Eosin dye

The whole lung specimens from the pulmonary metastasis mouse model were paraffin-embedded and sliced into tissue sections with a thickness of 3–4 μm. After dewaxing, the tissue sections were stained with hematoxylin dye (Servicebio) for 3–8 min, differentiated with 1% hydrochloric acid alcohol, and turned blue with 0.6% aqueous ammonia to complete nuclear staining, and cytoplasmic staining was performed by immersing sections in eosin dye (Servicebio) for 1–3 min. After dehydration and drying, the tissue sections were sealed with gum, digitally scanned, and analyzed.

Tissues fixed in 4 % paraformaldehyde (Beyotime, China) were dehydrated and embedded to form tissue wax blocks, which were cut into slices with a thickness of 4 μm with a paraffin microtome (Leica, Germany). Paraffin sections were dipped into xylene twice for 20 min each, anhydrous ethanol twice for 5 min each, and 75 % ethanol to deparaffinize and rinse with tap water. Then, the sections were treated with haematoxylin differentiation solution (Servicebio, China) and rinsed with tap water. Following immersion in 85 % ethanol for 5 min and 95 % ethanol for 5 min, the sections were placed in eosin dye (Servicebio, China) for 5 min and then dehydrated. Finally, the slides were sealed with neutral gum (SCRC, China), observed and photographed under a microscope (Olympus, Japan).

The paraffin sections were dewaxed in water, and stained with hematoxylin dye solution (Servicebio, Wuhan, China) for 3–5 min. Then, these were washed by tap water, differentiated by differentiation solution (Servicebio, Wuhan, China), washed again by tap water, colored blue by the blue solution (Servicebio, Wuhan, China), and washed by water again. Afterwards, the sections were successively dehydrated with 85 and 95% gradient alcohol for 5 min, respectively, and stained with eosin dye (Servicebio, Wuhan, China) for 5 min. Finally, the tablet was dehydrated and sealed.