The PAXgene Saliva Collector is part of a comprehensive pre-analytical workflow for human saliva collection, stabilization, transport and storage through nucleic acid extraction and analyses. The collection device contains a stabilizing solution which stabilizes the DNA levels in human saliva samples by protecting DNA from degradation and inhibiting bacterial growth over time. It is compatible with existing QIAGEN automated extraction kits, such as the QIAsymphony DNA Midi Kit on the QIAsymphony SP Instrument and the QIAamp DNA Mini Kit on the QIAcube (Classic and Connect), as well as for manual extraction with the QIAamp DNA Mini and Gentra Puregene Cell Kits.
In addition to DNA stabilization, SARS-CoV-2 (severe acute respiratory syndrome coronavirus type 2)-derived RNA copy numbers are preserved when saliva is collected in the PAXgene Saliva Collector. The viral RNA can be prepared with the QIAamp Viral RNA Mini Kit and quantified with QuantiTect Probe RT-PCR on QIAGEN Rotor-Gene® Q (See " PAXgene Saliva workflow").
Samples collected with the PAXgene Saliva Collector have stable DNA levels for up to 24 months at temperatures up to 25°C. This longevity is possible as DNA is preserved from chemical and enzymatic degradation and bacterial growth is inhibited. After 24 months of storage DNA still retains both high yield and quality if isolated with the QIAGEN DNA extraction kits and protocols specified (See " DNA yield, purity, and integrity is maintained after 24-month storage of stabilized saliva in PAXgene Saliva Collector"). Long-term storage of stabilized saliva at −20 or −80°C is possible (storage study is ongoing).
Protocols and supplementary protocols (see resource section) provide a broad range of existing, proven and flexible QIAGEN DNA extraction solutions for throughput and sample input volume. These protocols provide DNA that is high yield and purity, free of PCR inhibitors.
For manual extraction using silica membrane technology, supplementary protocols are available for the QIAamp DNA Mini Kit spin and vacuum protocols. This kit can also be used for automated DNA extraction on the QIAcube (classic) and QIAcube Connect (See "Automated DNA extraction with QIAamp DNA Mini Kit on the QIAcube (classic) and QIAcube Connect"). Standard protocols for the QIAcube (classic) and QIAcube Connect are available on www.QIAGEN.com. To download a QIAcube protocol sheet, select “DNA” as the application, “QIAamp DNA Mini” for the kit, and “PAXgene Saliva Collector” as the starting material. Optimized protocols for the QIAamp DNA Mini Kit provide high yields of pure DNA, and flexible elution volumes allow DNA concentration to be easily optimized for each downstream application.
The Puregene procedure uses a modified salting-out precipitation method for purification of DNA. The supplementary protocol “Protocol “Purification of DNA from the PAXgene® Saliva Collectors using the Gentra® Puregene® Cell Kit” for the Gentra Puregene Cell Kit offers a simple solution for manual DNA extraction from different sample input volumes, including the whole content of a PAXgene Saliva Collector (See " Consistently high DNA yield and purity with flexible sample input volume when using the Gentra Puregene Cell Kit").
Medium- to high-throughput automated purification of DNA from the PAXgene Saliva Collector is possible on the QIAsymphony SP Instrument using protocols for the QIAsymphony DNA Midi Kit for sample volumes of 400 and 1000 µl. High reproducibility is guaranteed through standardized magnetic bead processing technology (See " Automated DNA extraction using the QIAsymphony DNA Midi Kit on the QIAsymphony SP Instrument.").
Stabilization characteristics have been shown to be robust against elevated levels of endogenous and potentially interfering substances, under- or overfilling and inappropriate mixing (see technical note ´PAXgene Saliva Collector Robustness Study´). DNA isolated with the specified QIAGEN DNA extraction kits and protocols is free from PCR inhibition and is compatible with downstream analytical PCR assays, including digital PCR with the QIAGEN QIAcuity Digital PCR system (See " Absolute Quantification of Gene Copies by dPCR using the QIAcuity Digital PCR system"), and next-generation sequencing (NGS) molecular test methods.
In addition to the genomic DNA, the SARS-CoV-2 (severe acute respiratory syndrome coronavirus type 2) is stabilized in saliva collected with the PAXgene Saliva Collector for at least 4 days (96 hours) at 20°C. The genome of SARS-CoV-2 consists of RNA and can be isolated from the saliva with QIAamp Viral RNA Mini Kit for quantification with QuantiTect Probe RT-PCR and SARS-CoV-2 N1+N2 Assay Kits on QIAGEN Rotor-Gene Q ( See " PAXgene Saliva stabilizing solution maintains SARS-CoV-2 derived RNA levels for at least 96 h and reduces infectivity of SARS-CoV-2" part A).
While the PAXgene Saliva Collector stabilizes SARS-CoV-2 particles for detection and quantification it efficiently inactivates the virus, rendering it unable to infect and replicate in a cell culture model (See " PAXgene Saliva stabilizing solution maintains SARS-CoV-2 derived RNA levels for at least 96 h and reduces infectivity of SARS-CoV-2" parts B, C, D).