TY - JOUR
T1 - Preparation of single-cell suspensions from the human placenta
AU - Garcia-Flores, Valeria
AU - Xu, Yi
AU - Pusod, Errile
AU - Romero, Roberto
AU - Pique-Regi, Roger
AU - Gomez-Lopez, Nardhy
N1 - Publisher Copyright:
© 2022, Springer Nature Limited.
PY - 2023/3
Y1 - 2023/3
N2 - Single-cell RNA-sequencing (scRNA-seq) allows the characterization of cellular composition and interactions in complex tissues. An essential prerequisite for scRNA-seq is the preparation of high-quality single-cell suspensions. So far, no protocols have been described for preparing such suspensions from the placenta, an essential organ for fetal development and a site of maternal–fetal immune interaction. Here we describe a protocol for the preparation of high-quality single-cell suspensions from human placental tissues—namely, the basal plate, placental villi and chorioamniotic membranes. The protocol outlines the collection of tissues from the placenta, tailored dissociation procedures for each tissue, and the cryopreservation of single-cell suspensions for multiplex sequencing library preparation. The protocol can be performed by a qualified investigator with basic working knowledge of placental structure. Moreover, the single-cell suspensions generated by using this protocol are compatible with droplet-based scRNA-seq technology, such as the 10x Genomics Chromium system. This protocol reliably produces single-cell suspensions from the placental tissues with high yield and viability for scRNA-seq. This protocol takes ~6 h to complete from tissue collection to cryopreservation of single-cell suspensions, and an additional 2 h for thawing of cryopreserved single cells.
AB - Single-cell RNA-sequencing (scRNA-seq) allows the characterization of cellular composition and interactions in complex tissues. An essential prerequisite for scRNA-seq is the preparation of high-quality single-cell suspensions. So far, no protocols have been described for preparing such suspensions from the placenta, an essential organ for fetal development and a site of maternal–fetal immune interaction. Here we describe a protocol for the preparation of high-quality single-cell suspensions from human placental tissues—namely, the basal plate, placental villi and chorioamniotic membranes. The protocol outlines the collection of tissues from the placenta, tailored dissociation procedures for each tissue, and the cryopreservation of single-cell suspensions for multiplex sequencing library preparation. The protocol can be performed by a qualified investigator with basic working knowledge of placental structure. Moreover, the single-cell suspensions generated by using this protocol are compatible with droplet-based scRNA-seq technology, such as the 10x Genomics Chromium system. This protocol reliably produces single-cell suspensions from the placental tissues with high yield and viability for scRNA-seq. This protocol takes ~6 h to complete from tissue collection to cryopreservation of single-cell suspensions, and an additional 2 h for thawing of cryopreserved single cells.
UR - http://www.scopus.com/inward/record.url?scp=85139383862&partnerID=8YFLogxK
U2 - 10.1038/s41596-022-00772-w
DO - 10.1038/s41596-022-00772-w
M3 - Article
C2 - 36451054
AN - SCOPUS:85139383862
SN - 1754-2189
VL - 18
SP - 732
EP - 754
JO - Nature Protocols
JF - Nature Protocols
IS - 3
ER -