Allplex HPV HR Detection assay fulfils all clinical performance and reproducibility validation requirements for primary cervical cancer screening.

Human papillomavirus (HPV)-based screening offers better protection against cervical cancer compared to cytology, but HPV screening assays must adhere to validation requirements of the international guidelines to ensure optimal performance. Allplex HPV HR Detection (Allplex) assay, launched in the late 2022, is a fully automated real-time PCR-based assay utilizing innovative technology that enables quantification and concurrent distinction of 14 high-risk HPV genotypes (HPV16,18,31,33,35,39,45,51,52,56,58,59,66 and 68). We assessed the validity of the Allplex for cervical cancer screening purposes, via comparison to a clinically validated comparator assay (Hybrid Capture 2; HC2), and through assessment of intra-laboratory reproducibility and inter-laboratory agreement. A clinical validation panel comprised of 973 residual ThinPrep samples was obtained from women aged 30-64 years participating in the organized Slovenian screening program, of these 863 were from women undergoing their regular screening visit after a previous negative screen test while 110 were from women with underlying cervical intraepithelial neoplasia grade 2 or worse (CIN2+) lesions. The Allplex's relative clinical sensitivity for detection of CIN2+ and CIN3+ were 1.01 (95%CI;0.98-1.04) and 0.98 (95%CI;0.95-1.02), compared to that of HC2. At recommended thresholds of ≥98% and ≥90%, the Allplex's clinical sensitivity and specificity (p=0.0004 and p=0.02, respectively) were non-inferior to HC2. High intra-laboratory reproducibility and inter-laboratory agreement, both overall (98.1% and 97.9%, respectively) and at genotype level (>98.7%) was observed. In addition, analytical genotype-specific performance of Allplex was compared to that of its predecessor Anyplex HPV HR; high overall agreement was observed (96.3%; kappa value 0.88), with some variations in performance. In conclusion, Allplex met all validation criteria described in the international guidelines on sensitivity, specificity and laboratory reproducibility and can be considered clinically validated for primary cervical cancer screening.
Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: AOV has received reimbursement of travel expenses for attending conferences and honoraria for speaking from Abbott Molecular, Qiagen and Seegene. MP's and AOV's institution received research funding, free-of-charge reagents, and consumables to support research in the last 3 years from Qiagen, Seegene, Abbott, and Roche, all paid to their employer. KC & LC's institution received research funding, free-of-charge reagents, and consumables to support research in the last three years from Cepheid, Euroimmun, GeneFirst, Self-screen, Hiantis, Seegene, Roche, Abbott, Hologic, Vaccitech and Daye, all paid to their employer. AZ and ŠS declare no conflicts of interest. This research was supported by Seegene. Seegene had no role in the study design, data Agency (grant no. P3-00083). collection, analysis and interpretation of the data, manuscript preparation and the decision to publish present manuscript. AOV and MP are supported by the Horizon 2020 Framework Program for Research and Innovation of the European Commission, through the RISCC Network (grant no. 847845) and by the Slovenian Research
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