Anti vimentin

Fresh tissues were immobilized with 4% formalin for 48 h at 37°C and subsequently washed with cool phosphate-buffered saline (PBS) three times for 5 min each time. Tissue samples were dehydrated using ethanol, embedded in paraffin, and then cut them into 4-μm-thick sections of 2 mm diameter to produce TMAs. The paraffin sections were dewaxed in an incubator at 65°C overnight, before being deparaffinized in xylene and rehydrated using a decreasing ethanol gradient. Sections were placed in ethylene diamine tetraacetic acid (cat.no. 0085; Beyotime) and heated in a microwave for 10 min for antigen repair. A 3% hydrogen peroxide was used to eliminate endogenous peroxidase at 37°C for 30 min. Sections were incubated with anti-HNRNPC (dilution 1:50, cat. no. ET1611-2; HUABIO), anti-ki67 (dilution 1:10000, cat. no. 27309; Proteintech), anti-CD34 (dilution 1:50, cat. no. ET1606-11; HUABIO), anti-Vimentin (dilution 1:100, cat. no. ET1610-39; HUABIO), and anti-E-Cadherin (dilution 1:50, cat. no. ET1607-75; HUABIO) antibodies at room temperature overnight, and then with secondary antibody (dilution: 1:1, cat.no. K5007; Dako) for 1 h at room temperature. Finally, sections were stained with an IHC kit (cat. no. K5007; Dako) according to the manufacturer’s protocol.