| Summary: | <p>Abstract</p> <p>Background</p> <p>For titer assessment of human herpesvirus 6 (HHV-6), IFA targeting viral proteins or a TCID<sub>50</sub> method with ocular inspection for CPE can be used. These methods rely on the subjective decision of the assessor, obstructing the ability to obtain unanimous results.</p> <p>Findings</p> <p>We have developed and validated an alternative TCID<sub>50</sub> read-out approach where infection in the titration culture plate is assessed by viral DNA load change by quantitative PCR. A ten time increase in viral DNA load was determined as cut point for infection since that yielded a maximum correlation with viral protein expression (93%). The average intra-assay CV was 9% for quantitative PCR read-out of TCID<sub>50</sub> compared to 45% for ocular inspection read-out of TCID<sub>50</sub>, 14% for IFA read-out of TCID<sub>50</sub>, and 43% for an infectious units approach using IFA. The average inter-assay CV for quantitative PCR read-out of TCID<sub>50</sub> was 73%, compared to 66%, 25% and 77% for the ocular inspection read-out for TCID<sub>50</sub>, IFA read-out of TCID<sub>50</sub> and infectious unit approaches respectively.</p> <p>Conclusions</p> <p>The quantitative PCR based read-out of TCID<sub>50</sub> proved to be more robust and easier to interpret than traditional TCID<sub>50</sub> assessment approaches for HHV-6, and therefore it might be considered as an alternative method.</p>
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