title: Toward Rare Blood Cell Preservation for RNA Sequencing.
abstract: Cancer is driven by various events leading to cell differentiation and disease progression. Molecular tools are powerful approaches for describing how and why these events occur. With the growing field of next-generation DNA sequencing, there is an increasing need for high-quality nucleic acids derived from human cells and tissues-a prerequisite for successful cell profiling. Although advances in RNA preservation have been made, some of the largest biobanks still do not employ RNA blood preservation as standard because of limitations in low blood-input volume and RNA stability over the whole gene body. Therefore, we have developed a robust protocol for blood preservation and long-term storage while maintaining RNA integrity. Furthermore, we explored the possibility of using the protocol for preserving rare cell samples, such as circulating tumor cells. The results of our study confirmed that gene expression was not impacted by the preservation procedure (r(2) > 0.88) or by long-term storage (r(2) = 0.95), with RNA integrity number values averaging over 8. Similarly, cell surface antigens were still available for antibody selection (r(2) = 0.95). Lastly, data mining for fusion events showed that it was possible to detect rare tumor cells among a background of other cells present in blood irrespective of fixation. Thus, the developed protocol would be suitable for rare blood cell preservation followed by RNA sequencing analysis.
authors.affiliation: Science for Life Laboratory, Royal Institute of Technology (KTH), Stockholm, Sweden.
authors.forename: Sanja
authors.initials: S
authors.lastname: Vickovic
authors.affiliation: Science for Life Laboratory, Royal Institute of Technology (KTH), Stockholm, Sweden.
authors.forename: Afshin
authors.initials: A
authors.lastname: Ahmadian
authors.affiliation: Department of Oncology-Pathology, Karolinska Biomics Center, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Rolf
authors.initials: R
authors.lastname: Lewensohn
authors.affiliation: Science for Life Laboratory, Royal Institute of Technology (KTH), Stockholm, Sweden. Electronic address: joakim.lundeberg@scilifelab.se.
authors.forename: Joakim
authors.initials: J
authors.lastname: Lundeberg
mesh_terms: D001773:Blood Cells / Q000378:metabolism*; D064368:Healthy Volunteers; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D008297:Male; D009369:Neoplasms / Q000235:genetics*; D011309:Preservation, Biological / Q000379:methods*; D012313:RNA / Q000235:genetics / Q000302:isolation & purification*; D017423:Sequence Analysis, RNA; D013048:Specimen Handling / Q000379:methods*; D014407:Tumor Cells, Cultured
chemical_list: D012313:RNA