Dataset Information

An optimised saliva collection method to produce high-yield, high-quality RNA for translational research.

ABSTRACT: Saliva represents an ideal matrix for diagnostic biomarker development as it is readily available and requires no invasive collection procedures. However, salivary RNA is labile and rapidly degrades. Previous attempts to isolate RNA from saliva have yielded poor quality and low concentrations. Here we compare collection and processing methods and propose an approach for future studies. The effects of RNA stabilisers, storage temperatures, length of storage and fasting windows were investigated on pooled saliva samples from healthy volunteers. Isolated RNA was assessed for concentration and quality. Bacterial growth was investigated through RT-PCR using bacterial and human primers. Optimal conditions were implemented and quality controlled in a clinical setting. The addition of RNAlater increased mean RNA yield from 4912 ng/?l to 15,473 ng and RNA Integrity Number (RIN) from 4.5 to 7.0. No significant changes to RNA yield were observed for storage at room temperature beyond 1 day or at -80 °C. Bacterial growth did not occur in samples stored at ambient temperature for up to a week. There was a trend towards higher RNA concentration when saliva was collected after overnight fasting but no effect on RIN. In the clinic, RNA yields of 6307 ng and RINs of 3.9 were achieved, improving on previous reports. The method we describe here is a robust, clinically feasible saliva collection method using preservative that gives high concentrations and improved RINs compared to saliva collected without preservative.

SUBMITTER: Sullivan R

PROVIDER: S-EPMC7062242 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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An optimised saliva collection method to produce high-yield, high-quality RNA for translational research.

Sullivan Roisin R Heavey Susan S Graham David G DG Wellman Rachel R Khan Saif S Thrumurthy Sri S Simpson Benjamin S BS Baker Tina T Jevons Sarah S Ariza Jose J Eneh Victor V Pye Hayley H Luxton Hayley H Hamoudi Rifat R Whitaker Hayley H Lovat Laurence B LB

PloS one 20200309 3

Saliva represents an ideal matrix for diagnostic biomarker development as it is readily available and requires no invasive collection procedures. However, salivary RNA is labile and rapidly degrades. Previous attempts to isolate RNA from saliva have yielded poor quality and low concentrations. Here we compare collection and processing methods and propose an approach for future studies. The effects of RNA stabilisers, storage temperatures, length of storage and fasting windows were investigated o ...[more]

PMID: 32150588

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Project description:Systematic collection of fresh tissues for research at the time of diagnostic image-guided breast biopsy has the potential to fuel a wide variety of innovative studies. Here we report the initial experience, including safety, feasibility and laboratory proof-of-principle, with the collection and analysis of research specimens obtained via breast core needle biopsy immediately following routine clinical biopsy at a single institution over a 14-month period. Study eligibility criteria consisted of patients age 18 or older who were undergoing a clinical breast needle biopsy. Patients providing informed consent underwent one or two additional core biopsies following collection of all necessary clinical specimens. In total, 395 patients were approached and 270 consented to the research study, yielding a 68.4% consent rate. Among consenting patients, 238 lesions were biopsied for research, resulting in 446 research specimens collected. No immediate complications were observed. During the same period, zero complications were reported among the non-research procedures (1622 breast biopsy procedures). Representative research core specimens showed high diagnostic concordance with clinical core biopsies. Flow cytometry demonstrated consistent recovery of hundreds to thousands of viable cells per research core. Among a group of HER2+ tumor research specimens, HER2 assessment by flow cytometry correlated highly with immunohistochemistry (IHC) staining, and in addition revealed extensive inter- and intra-tumoral variation in HER2 levels of potential clinical relevance. Suitability for single-cell transcriptomic analysis was demonstrated for a triple-negative tumor core biopsy, revealing substantial cellular diversity in the tumor immune microenvironment, including a prognostically relevant T cell subpopulation. Thus, collection of fresh tissues for research purposes at the time of diagnostic breast biopsy is safe, feasible and efficient, and may provide a high-yield mechanism to generate a rich tissue repository for a wide variety of cross-disciplinary research.

Project description:BackgroundThe ubiquitous use of mobile phones in sending and receiving text messages has become a norm for young people. Undeniably, text messaging has become a new and important communication medium not only in the social realm but in education as well. The aim of this study is to evaluate the effectiveness of using text messaging as a means to collect data for a medical research project.A cross sectional study was carried out during a double blind, randomized controlled trial to assess the efficacy and safety of a probiotic in the management of Irritable Bowel Syndrome (IBS). The study aim was to assess the response rate of weekly symptom reports via Short Message Service (SMS). The subjects were undergraduates in a private medical university in Malaysia. They were identified through a previous university wide study as suffering from IBS based on Rome III criteria. The subjects were randomly assigned to either the treatment arm receiving a daily probiotic, or the placebo arm. They were required to score their symptoms using eight-item-questionnaires at baseline, and thereafter weekly, for a total of 8 weeks. All subjects were given the choice to communicate their symptom scores by text messaging via mobile phones or by email. SMS text messages were sent to remind trial subjects to attend face-to-face visits and to complete a paper based 34-item-questionnaires on IBS quality of life assessment at baseline and at end of 8 weeks.FindingsThe response rate of weekly symptom scores via Short Message Service (SMS) from a total of 38 subjects was 100%. Through the study, 342 reports were submitted: 33.3% of these were received on the due date without reminder, 60.0% one day after the deadline, after a single reminder, 6.1% 2-3 days after the deadline, after 2-3 reminders and 0.6% 5 days after the deadline, after SMS, phone reminder and face-to-face encounter. All SMS symptom reports, whether on time or late, were complete. With the help of SMS reminder, all trial subjects completed the paper based IBS quality of life assessment at baseline and at end of study.ConclusionsThis study found using text messaging via mobile phone an excellent instrument for collecting weekly symptom reports in response to trial medication, reminding trial subjects to attend face to face visits and completing more complex paper based evaluation. The 100% response rate of weekly symptom reports was facilitated by using simple number codes for SMS submission.Trial registrationNot appropriate.

Dataset Information

An optimised saliva collection method to produce high-yield, high-quality RNA for translational research.

Publications

An optimised saliva collection method to produce high-yield, high-quality RNA for translational research.

Similar Datasets

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