BACKGROUND Xenotropic murine leukemia virus-related virus (XMRV) is a novel human gammaretrovirus that was first identified in patients with prostate cancer in 2006. Subsequent studies have shown that XMRV is also detected in patients with chronic fatigue syndrome (CFS) and even in some healthy controls and blood donors. However, some conflicting findings have been reported by different laboratories or in different regions. The association of XMRV with human diseases and the prevalence of XMRV in different populations needs to be further determined. METHODS XMRV was screened in 391 blood samples from healthy blood donors in China. Nested reverse transcription-polymerase chain reaction (PCR) was used to amplify gag and env genes of XMRV from total RNA of peripheral blood mononuclear cells (PBMNCs) and plasma, respectively. Quantitative PCR was performed to detect XMRV env gene in genomic DNA of PBMNCs. To enhance the detection sensitivity, plasma was added into LNCaP cells to amplify XMRV in the plasma samples. RESULTS No XMRV was found in the 391 blood donors in China or in the LNCaP cells inoculated with plasma from the blood donors. CONCLUSIONS Both PCR and virus isolation in highly permissive LNCaP cells failed to detect XMRV in 391 Chinese blood donors, indicating that XMRV infection might not be present in blood donors in China.