RNeasy Protect Saliva Mini Kit
For stabilization and purification of total RNA from human saliva
The RNeasy Protect Saliva Mini Kit includes the RNAprotect Saliva Reagent, which stabilizes RNA in saliva samples to preserve the gene expression profile, and the RNeasy Micro Kit, which purifies and concentrates total RNA using specialized RNeasy MinElute spin columns. Stabilized saliva samples can be conveniently stored or shipped at 37°C (1 day), 15–25°C (14 days), or 2–8°C (4 weeks) before RNA purification. RNA is purified from 200 μl saliva and eluted in a volume of just 14 μl.
Saliva stabilized with RNAprotect Saliva Reagent can be stored or transported for up to 1 day at 37ºC, up to 14 days at room temperature (15–25ºC), or up to 4 weeks at 2–8ºC, or can be archived at –20ºC or –80ºC (see figure "Effective stabilization of β-actin transcript"). Purified RNA from saliva, as an alternative to blood, allows microarray analysis for research into oral and systemic diseases, and gene expression analysis of RNA biomarkers for cancer by real-time RT-PCR (see figures "Minimization of transcript degradation in saliva" and "Stable mRNA levels of cancer biomarkers").
The RNeasy Protect Saliva Mini Kit provides a complete RNA protection and isolation solution, from sample harvesting to pure RNA, in one kit. Proven RNeasy technology, combined with the RNA-stabilizing properties of RNAprotect Saliva Reagent, allows purification of high-quality, intact RNA. Saliva contains bacteria in addition to cells and free-circulating nucleic acids of human origin. When saliva is collected from a donor and mixed with RNAprotect Saliva Reagent, the RNA is immediately stabilized and uncontrolled growth of bacteria is prevented, ensuring reliable results in downstream gene expression analysis. Following stabilization, RNeasy technology simplifies total RNA isolation by combining the stringency of guanidine-isothiocyanate lysis with the speed and purity of silica-membrane purification. RNA in bacteria is already degrading prior to collection and stabilization in RNAprotect Saliva Reagent, and will be of lower quality than RNA purified from cultured cells and animal tissues.
Saliva (200 µl) collected from a donor is immediately mixed with RNAprotect Saliva Reagent to stabilize the RNA in the saliva. Total RNA is purified from stabilized saliva using the RNeasy Micro Kit (see flowchart "RNeasy Protect Saliva procedure"). The saliva sample is first centrifuged, and Buffer RLT is then added to the resulting pellet. After addition of ethanol, the sample is applied to an RNeasy MinElute spin column, where RNA binds to the membrane after centrifugation. Traces of genomic DNA are removed by DNase digestion, and contaminants are washed away in several wash steps. Highly pure RNA is then eluted using RNase-free water in a volume of just 14 µl. RNAprotect Saliva Reagent also stabilizes DNA. Protocols for purification of DNA, or RNA and DNA, from stabilized saliva are available.
Total RNA purified from stabilized saliva using the RNeasy Micro Kit represents a noninvasive alternative to blood analysis. The kit allows research into the identification of RNA biomarkers for:
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