Eliteingenius sp 200

SARS-CoV-2 RNA was searched in all the cases on sections from olfactory bulb and tract, postrolandic cortex and rostral medulla as previously described (10). FFPE 10 micron sections were pre-treated using 160 µL Deparaffinization Solution (Qiagen, Hilden, Germany) with 180 μL of tissue lysis buffer (ATL buffer, Qiagen) and 20 μL of protease (Proteinase K solution, Qiagen, Germany). Tissues were incubated overnight at 56 °C and one-hour at 90 °C. Then extraction and amplification of nucleic acids were performed using ELITeInGenius^® instrument with ELITeInGenius SP 200 and SARS-CoV-2 ELITe MGB Kit (ELITechGroup, Milano, Italy). The tissue viral load was reported as number of copies/microgram RNA. Quantification of RNA was performed using Qubit Assay (Invitrogen by ThermoFisher). Positive results below the lower limit of quantification (2 copies/reaction) were reported as <15 copies/microgram RNA. The identification of key mutations in SARS-CoV-2 Spike Protein gene (E484K, N501Y, HV69-70 del, L452R, K417T, K417N, W152C) was performed with Allplex SARS-CoV-2 Variants I and II Assay, Seegene.

For the automatic extraction with the ELITe InGenius® SP200 (ELITechGroup) system of SARS-CoV-2 RNA, 200 μL of the medium from UTM® tubes containing 3 mL was used, following the manufacturer's instructions. Samples were eluted in 100 μL elution buffer and used as the template for downstream analysis. Extracted RNA was stored at −80°C.

For each SARS-CoV-2 pharyngo-nasal swab positive patient, two samples from medullary and cortical regions of the retrieved ovarian tissue were cut and sent together with peritoneal swabs and washings to the virology department for SARS-CoV-2 RNA detection as previously reported by Seracchioli et al. (10 (link)).
Briefly, extraction and purification of nucleic acids were performed using ELITe InGenius^® instrument with the ELITe InGenius SP 200 (ELITechGroup, Puteaux, France). For the qualitative detection of SARS-CoV-2 the GeneFinderTM COVID-19 PLUS RealAmp Kit (OSANGHealthcare, Gyeonggi, South Korea) was used on the ELITe InGenius^® instrument. The kit detects RNA-dependent RNApolymerase (RdRp), envelope (E), and nucleocapsid (N) genes of SARS-CoV-2.