Detection of XMRV RNA and DNA in viral Ab-positive samples. (A) RNA was purified from 1 mL of coculture supernatant of activated PBMCs and LNCap-FGC cells (lane 2) or 1 ml plasma (lanes 3-6). For one-step RT-PCR, 15 μl of 60 μl eluted RNA was amplified in a 25 μl volume. CFS patients C4 and C32 tested positive for XMRV Abs but C1 was negative. (B) Detection of XMRV env by TaqMan real-time PCR assay. Duplicated test samples of diluted XMRV plasmid (VP62) were amplified. The detection limit of the TaqMan real-time PCR was 4 copies/reaction determined by VP62 plasmid. (C) Duplicated test samples without template DNA in negative control (N) or with genomic DNA extracted from PBMCs of a viral Ab-positive PC patient (P24) and healthy volunteers (HV) were amplified as for (B).