DMSO cryopreservation is the method of choice to preserve cells for droplet-based single-cell RNA sequencing.
Sci Rep
; 9(1): 10699, 2019 07 23.
Article
em En
| MEDLINE
| ID: mdl-31337793
ABSTRACT
Combining single-cell RNA sequencing (scRNA-seq) with upstream cell preservation procedures such as cryopreservation or methanol fixation has recently become more common. By separating cell handling and preparation, from downstream library generation, scRNA-seq workflows are more flexible and manageable. However, the inherent transcriptomic changes associated with cell preservation and how they may bias further downstream analysis remain unknown. Here, we present a side-by-side droplet-based scRNA-seq analysis, comparing the gold standard - fresh cells - to three different cell preservation workflows dimethyl sulfoxide based cryopreservation, methanol fixation and CellCover reagent. Cryopreservation proved to be the most robust protocol, maximizing both cell integrity and low background ambient RNA. Importantly, gene expression profiles from fresh cells correlated most with those of cryopreserved cells. Such similarities were consistently observed across the tested cell lines (R ≥ 0.97), monocyte-derived macrophages (R = 0.97) and immune cells (R = 0.99). In contrast, both methanol fixation and CellCover preservation showed an increased ambient RNA background and an overall lower gene expression correlation to fresh cells. Thus, our results demonstrate the superiority of cryopreservation over other cell preservation methods. We expect our comparative study to provide single-cell omics researchers invaluable support when integrating cell preservation into their scRNA-seq studies.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Criopreservação
/
Dimetil Sulfóxido
/
Análise de Sequência de RNA
/
Análise de Célula Única
Limite:
Humans
Idioma:
En
Revista:
Sci Rep
Ano de publicação:
2019
Tipo de documento:
Article
País de afiliação:
Alemanha