POLYMERASE-CHAIN-REACTION BASED ASSESSMENT OF LEUKOREDUCTION EFFICACYUSING A CYTOMEGALOVIRUS DNA TRANSFECTED HUMAN T-CELL LINE

Background and Objectives: Detection of CMV DNA by PCR in seropositive blood donors is affected by nucleic acid extraction, amplification co nditions and PCR product detection sensitivity. Clinical studies have shown that leukoreduced blood products are as effective as CMV-seroneg ative blood products in minimizing transfusion-associated CMV infectio n. We developed a PCR-based test model to assess the efficacy of leuko reduction on the removal of CMV DNA from blood donations, Materials an d Methods: Whole blood units were spiked with a human Jurkat T-lymphoc yte cell line which had been stably transfected with a CMV DNA immedia te early sequence. All blood units were either CMV seronegative or sho wn to be CMV PCR negative. The amount of CMV DNA by PCR before, during and after filtration of blood units with third-generation leukocyte r eduction filters were determined using a semi-quantitative adaptation of the Digene SHARP Signal(TM) System Assay (DSSSA). Results. Whole bl ood units spiked with 1-2 x 10(6) CMV transfected Jurkat cells contain ed no detectable CMV DNA after leukoreduction. With a higher inoculum of 2.9 x 10(8) transfected Jurkat cells per unit, CMV DNA was detected after leukoreduction; however, there was an approximate 3 log decreas e in the amount of CMV DNA detected. Conclusion: This CMV DNA transfec ted human Jurkat cell line in conjunction with semi-quantitative CMV P CR may be used to model leukoreduction efficacy and provides evidence that leukoreduction of blood products can decrease the amount of cellu lar CMV DNA by approximately 3 logs. (C) 1998 Elsevier Science B.V. Al l rights reserved.