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The reaction of β-catenin, PCNA, or Ki67 antibody (Abcam) took place for 1 h at room temperature, followed by incubation by biotin-labeled secondary antibodies, and the slides were then developed using 3,3′-diaminobenzidine (DAB) solution and counterstained with hematoxylin staining (BASO, Shanghai, China).

Finally, each sample was resuspended to a volume of 500 μL with 1× binding buffer and was analyzed through flow cytometry within 30 min.

More than one-third of RCC patients present at advanced stages, and nearly half of the patients who experience curative therapies such as surgical resection suffer recurrence in the subsequent follow-up.

Therefore, it is essential to find the molecular mechanism of tumor cell metastasis in primary lesions, which can improve the treatment and prognosis.

It was found that Introduction Renal cell carcinoma (RCC), the most common type of adult kidney malignancy, accounts for 80% of all primary renal neoplasms and is composed of 2%–3% of all malignancies.

The occurrence of RCC has increased in recent years; worldwide, ~100,000 patients die of disease every year.

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