2.3 HCMV UL21.5 mRNA detection
To evaluate the presence of UL21.5 mRNA, nucleic acids were extracted
from 600µl of plasma using ELITe InGenius® SP1000. Realtime RT-PCR was
performed using CMV RNA ELITe MGB® kit (kindly provided by ELITech
Group) targeting the UL21.5 gene, according to the manufacturer’s
instructions. Nucleic acids extraction, UL21.5 cDNA synthesis and
amplification and results interpretation were performed on the ELITe
InGenius® platform (ELITech Group, USA). To obtain a reliable
quantification, standard samples (concentrated
102-104 copies/reaction) were tested
once every 60 days, following the manufacturer’s instructions. Results
were expressed in copies/ml. Samples in which UL21.5 mRNA was detected
but at levels below the quantification limit (30 copies/ml) were
assigned an arbitrary value of 10 copies/ml. Negative samples were
reported as “undetectable”.