Comparison of blood RNA isolation methods from samples stabilized in Tempus tubes and stored at a large human biobank

Additional file 3. Evaluation of reference gene stability. The non-normalized raw Cq-values of reference small ncRNAs for adult (n=12 Tempus tubes) and cord blood (n=3 Tempus tubes) samples collected in the Tempus tubes of the blood-RNA samples isolated using the three RNA isolation protocols

Long-term storage of blood RNA collected in RNA stabilizing Tempus tubes in a large biobank – evaluation of RNA quality and stability

Background: Establishing methods for secure long term storage of RNA is critical to realizing the promise of biobanks in biomedical research. Here, we describe the results of yearly analyses of the same set of umbilical cord and adult whole blood RNA collected in Tempus Blood RNA tubes and stored at -80°C, over a period of up to six years. We systematically investigated the effects of long-term storage of samples (75 Tempus tubes form three adult donors and 30 Tempus tubes from three cord blood donors) on the RNA quality and transcript stability of six selected genes (CDKN1A, FOS, IL1B, IL8, MYC and TP53). This is the first systematic study of both cord and adult blood samples stored for many years. Findings: The RNA purity and integrity, expressed as RIN-values, were stable up to six years of storage, and there were no storage-related deleterious effects on RNA purity. There were limited intra- and inter-individual variations in RNA yields; however, no consistent trend of decreasing RNA yield was observed with the duration of storage. Some long-term storage effects were found on the relative transcript levels of the six genes when compared to the year 0 samples. However, these changes were within ± 2–fold for both types of blood samples, except for two genes. Our results show that storage of these samples for up to six years did not have significant effects on the RNA quality and transcript stability of the six genes. Conclusions: Blood RNA is stable in Tempus tubes stored at -80°C over a period of six years. Intact and good-quality RNA suitable for transcript profiling analyses in epidemiological studies was obtained from blood samples stored in Tempus tubes. This suggests that blood samples collected in large biobanks–such as the Mother and Child (MoBa) Cohort at Norwegian Institute of Public Health (NIPH) and frozen in suitable collection tubes for total RNA stabilization, can be used for quantitative studies after at least six years of storage

MOESM1 of Comparison of blood RNA isolation methods from samples stabilized in Tempus tubes and stored at a large human biobank

Additional file 1. Target abundance of RNA transcripts. Evaluation of target abundance; i.e., whether low abundant RNA transcripts (RNA transcripts with Cq-values above 30 cycles) could be detected in blood-RNA samples isolated using the five blood-RNA isolation protocols. Low abundant mRNA and miRNA transcripts were detected in all analyzed samples. The average of non-normalized raw Cq-values of all targets from all adult (n=11-12 Tempus tubes) and cord blood (n=3-6 Tempus tubes) samples collected in the Tempus tubes of the blood-RNA samples isolated using the five RNA isolation protocols