Performance evaluation of Sanger sequencing for the diagnosis of primary XXXX and comparison with targeted next generation sequencing.

@entity1 @entity5201 @entity49814 @entity12297 @entity71531
Definitive diagnosis of primary @entity5201 (PH) currently utilizes sequential Sanger sequencing of the @entity12297 , GRPHR, and @entity49814 genes but efficacy is unproven. This analysis is time-consuming, relatively expensive, and delays in diagnosis and inappropriate treatment can occur if not pursued early in the diagnostic work-up. We reviewed testing outcomes of Sanger sequencing in 200 consecutive @entity1 samples referred for analysis. In addition, the Illumina Truseq custom amplicon system was evaluated for paralleled next-generation sequencing (NGS) of @entity12297 , @entity71531 , and @entity49814 in 90 known PH @entity1 . @entity12297 sequencing was requested in all @entity1 , permitting a diagnosis of PH1 in 50%. All remaining @entity1 underwent targeted exon sequencing of @entity71531 and @entity49814 with 8% diagnosed with PH2 and 8% with PH3. Complete sequencing of both @entity71531 and @entity49814 was not requested in 25% of @entity1 referred leaving their diagnosis in doubt. NGS analysis showed 98% agreement with Sanger sequencing and both approaches had 100% diagnostic specificity. Diagnostic sensitivity of Sanger sequencing was 98% and for NGS it was 97%. NGS has comparable diagnostic performance to Sanger sequencing for the diagnosis of PH and, if implemented, would screen for all forms of PH simultaneously ensuring prompt diagnosis at decreased cost.
