Anyplex 2 hpv28 kit

Cervicovaginal sample (5 mL) was self-collected by sub-study-1 individuals with the Delphi Screener® device (Delphi Bioscience, BV Scherpenzeel, the Netherlands) and transferred in a coded tube prior to sending to our laboratory within 24 h. DNA from 200 μL cell suspension was isolated with the MagNaPure 96 Total Nucleic Acids kit according to the manufacturer (Roche, Basel, Switzerland) and eluted in 100 μL elution buffer. Five μL was used for CT detection by real time PCR [14 (link)] and for genotyping of 28 HPV genotypes (6, 11, 16, 18, 26, 31, 33, 35, 39, 40, 42–45, 51–54, 56, 58, 59, 61, 66, 68–70, 73, and 82) by the Anyplex™ II HPV28 kit and the HPV51 RUO kit according to the manufacturer (Seegene, Seoul, South Korea). The HPV51 RUO kit was used for HPV51, which is otherwise not well covered with the Anyplex™ II HPV28 kit [15 (link)]. HPV- or CT-negative tests were valid only if their internal positive control was positive (human DNA target and spiked internal control, respectively). Negative controls were tested in parallel throughout the procedures to monitor contaminations.
DNA extraction and HPV genotyping (PGMY-CHUV assay) for sub-study-2 individuals have been described [16 (link), 17 (link)].

Two to four 3 μm-thick sections were cut from FFPE tissue samples and placed in Eppendorf Tube^® 1.5 mL. Subsequently, deparaffined using the MagCore^® Genomic DNA FFPE One-Step Kit (RBC Bioscience Corp. New Taipei City, Taiwan), following the manufacturer’s instructions (cartridge code: 405; execution time: 16 h; elution volume: 60 µL) using an extractor automated MagCore^® HF16 Plus nucleic acid (RBC Bioscience Corp.). Genotyping, viral load and co-infections were detected by Anyplex II HPV28 kit (Seegene, Seoul, Republic of Korea), which simultaneously identifies 28 genotypes: 19 high-risk HPV types (16, 18, 26, 31, 33, 35, 39, 45, 51, 52, 53, 56, 58, 59, 66, 68, 69, 73 and 82) and 9 low-risk HPV types (6, 11, 40, 42, 43, 44, 54, 61 and 70) by performing a multiplex PCR with CFX96 thermal cycler (Bio-Rad, Hercules, CA, USA). Data analysis and interpretation were automated with Seegene viewer software, according to the manufacturer’s instructions.

HPV testing is processed at the Molecular Diagnostic Laboratory of the Institute of Microbiology of CHUV. LBC residual samples are transferred from the Institute of Pathology to the Institute of Microbiology. DNA from 200 μL cell suspension is isolated with the MagNaPure 96 Total Nucleic Acids kit according to the manufacturer (Roche, Basel, Switzerland) and eluted in 100 μL elution buffer. Five μL are used for genotyping of 28 HPV genotypes (6, 11, 16, 18, 26, 31, 33, 35, 39, 40, 42–45, 51–54, 56, 58, 59, 61, 66, 68–70, 73, and 82) by the Anyplex™ II HPV28 kit according to the manufacturer (Seegene, Seoul, South Korea). HPV negative tests are valid only if their internal positive control is positive (human DNA target). Negative controls are tested in parallel throughout the procedures to monitor contaminations, as described in a previous publication [11 (link)].