Objective      To improve the method of primary culture of kidney cancer cells to get cells with high purity and similar phenotype.       Methods      Clinical specimens of renal clear cell carcinoma were digested and mechanically dissociated to prepare mono-cell suspension. The cells were subsequently cultured in ultra-low-adhesion plates

and tumor spheroids were formed after changing the medium to serum-free RPMI1640 medium (SFM) supplemented with mitogens. Tumor spheroids could be digested and passaged to get successive generation. Monolayer adherent cells (MACs) could be obtained from the digested tumor spheres and maintained as monolayer cultures in RPMI 1640 routine medium. Aliquots of the cell suspension were subjected to fluorescence-activated cell sorting to quantify the expression of cell surface antigen in the specimen cells and passaged cells.       Results      The improved method could dramatically increase the cells at high yield and high purity compared with traditional methods

and the cells obtained by serum-free culture had similar phenotype with primary cells and had no fibroblast cell incorporation.       Conclusion      The improved method is feasible and convenient and has high success rate. The cultured cells act as experimental model for research of renal cell carcinoma.