Multiplex Probe Amplification (MPA) technology dramatically expands multiplexing ability of a real-time PCR reaction. The technology allows detection of multiple targets in a single florescence channel using a mixture of hydrolysis based probes, partially complementary oligonucleotides and their melting curve analysis. The MPA technology was used to develop a quick, easy-to-use, sensitive and affordable genotyping assay that detects all 14 HR-HPV types in a single reaction – PapilloplexTM HR-HPV test.
In the present study, we carried out a comparative analysis of the performance of PapilloplexTM HR-HPV test with four well established assays on a panel of liquid based cytology (LBC) samples. Analytical specificity of the assay was also interrogated using the WHO HPV LabNet proficiency panel (2015).
The PapilloplexTM HR-HPV test was used to test 500 disease enriched cervical LBC samples obtained from the Scottish HPV Archive, Edinburgh with known concurrent pathology results. Samples were also tested by the Abbott rT HPV assay, the Qiagen Hybrid Capture 2 Assay, the Diamex Optiplex HPV Genotyping kit and Roche Linear Array HPV Genotyping test. Concordance between the comparator assays vs PapilloplexTM was performed using binomial test and McNemars test of proportions. As the samples were enriched for CIN2+ the applicability of clinical performance measures to screening settings are limited however these were assessed in terms of sensitivity and specificity for underlying CIN2+.
The PapilloplexTM assay was able to detect and genotype HPV HR types 16, 18, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66, 68a and 68b in a single reaction. The limit of detection for the assay was 5 genome copy numbers for HPV 16 and 18 in the WHO LabNet samples with 100% accuracy for genotyping. The overall proportional agreement of PapilloplexTM was high with Abbott rtHPV assay [95% (CI: 93 to 97%)], HC2 [90% (CI: 87-93%)], Linear Array [96% (94. 96%)] and Optiplex [94% (92, 96%)]. Type specific concordance was also high with all four assays.
These data indicate that the analytical performance of PapilloplexTM HR-HPV assay is comparable to established HPV assays at the level of generic HR-HPV detection and at the type specific level. The assay shows potential promise for both disease management and epidemiological applications. Further data on the clinical performance of the assay will be presented.