False-positive results of plasma PCR for cytomegalovirus DNA due to delayed sample preparation

Positive results by cytomegalovirus (CMV) PCR of plasma are considered pred ictive of active CMV infection in kidney allograft recipients. To assess wh ether contamination with leukocyte-derived CMV DNA can distort the results, aliquots of whole-blood samples from 60 CMV immunoglobulin G-positive pati ents with leukocyte CMV DNAemia were stored for up to 24 h at room temperat ure (RT) and at 4 degrees C before plasma preparation. Native and ultrafilt ered plasma samples were tested by CMV and beta-globin PCRs, Among 30 laten tly infected patients (negative for CMV pp65 antigens), low baseline rates (10%) and levels (median number of copies, 10 [per 10 mu l]) of CMV plasma DNAemia in native plasma samples increased significantly over time (after 4 h at RT, 37% [P < 0.001]; median number of copies, 45 [P < 0.001]). Simila r effects were found during storage at 4 degrees C. Ultrafiltration reduced the levels of CMV plasma DNAemia, but by 6 h of storage the Levels were si gnificantly elevated as well, CMV and beta-globin DNA kinetics in plasma we re parallel. In contrast, 30 actively infected patients (pp65 positive) had high baseline rates (87% in native samples) and levels (median number of c opies, 75) of CMV plasma DNAemia, No significant effects of storage or ultr afiltration and no concordance with beta-globin DNA kinetics were seen. In conclusion, delayed preparation of plasma samples bears a significant risk of false-positive CMV PCR results, probably due to leukocyte lysis, This ha s important implications in the clinical setting and for PCR standardizatio n.