Single-cell suspensions from the human placenta

A new protocol to support single-cell sequencing of the placenta will enable researchers to study the inner workings of this mysterious organ in unprecedented detail.

While the placenta is arguably one of the most important organs of the body, it is also one of the least well-understood. For the nine months of its existence, it carries out a host of activities that help protect and sustain the developing fetus – including facilitating the supply of oxygen and nutrients. Understanding the biology of the placenta is vital for improving the understanding of fetal development and the mechanisms of pregnancy complications, including preterm birth and pre-eclampsia.

Previous investigations of the placenta have largely been limited to detailed microscopic examination – or the molecular profiling of bulk tissue samples. But recent advances in single-cell technologies offer new opportunities to extract information at an individual cell level – enabling a much deeper examination of the cellular composition and interactions of this essential organ than ever before.

The first and most important step for single-cell applications is the isolation of individual cells from a tissue sample. But so far, no protocols have been described for preparing such suspensions from the placenta.

Step-by-step protocol

In a new paper published in Nature Protocols, researchers describe a method for the preparation of high-quality single-cell suspensions from human placental tissues – namely, the basal plate, placental villi, and chorioamniotic membranes.¹

The team set out a detailed protocol for the enzymatic dissociation of the placental tissues – along with guidance for quality control and single-cell applications. The distinct composition of placenta tissues precludes the use of a single digestion protocol, and so they validated two distinct methods tailored to the basal plate and placental villi, and chorioamniotic membranes.

The protocol can be performed by a qualified investigator with basic working knowledge of placental structure. It outlines the steps involved in the collection of tissues from the placenta, tailored dissociation protocols for each tissue, and the cryopreservation of single-cell suspensions for multiplex sequencing library preparation. It takes around six hours to complete from tissue collection to cryopreservation of single-cell suspensions, and an additional two hours for thawing of cryopreserved single cells.

The researchers include Type 1 ultrapure water generated from an ELGA PURELAB® Classic laboratory water purification system in the list of critical reagents for the protocol. They also state that using alternative reagents from other manufacturers may not guarantee the anticipated results.

Single-cell applications

This new protocol provides a useful systematic approach for the single-cell investigation of the human placenta, including the maternal-fetal interface. The new method reliably produces single-cell suspensions from the placental tissues with high yield and viability for single-cell sequencing.

Using single-cell sequencing data of the placenta, the researchers have contributed to the understanding of the maternal-fetal transmission of the coronavirus SARS-CoV-2, as well as to the investigation of maternal-fetal immune responses triggered by this virus.

The team hopes that the accessibility and detail of the protocol will encourage other investigators to pursue novel research questions in the field of maternal-fetal biology – uncovering the secrets of this underexplored organ.

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Reference:

1. Garcia-Flores V. et al. Preparation of single-cell suspensions from the human placenta. Nat. Protoc. (2022) https://doi.org/10.1038/s41596-022-00772-w/li>

Dr Alison Halliday

After completing an undergraduate degree in Biochemistry & Genetics at Sheffield University, Alison was awarded a PhD in Human Molecular Genetics at the University of Newcastle. She carried out five years as a Senior Postdoctoral Research Fellow at UCL, investigating the genes involved in childhood obesity syndrome. Moving into science communications, she spent ten years at Cancer Research UK engaging the public about the charity’s work. She now specialises in writing about research across the life sciences, medicine and health.