Feeder-free culture of naive human PSCs preserving embryonic, extraembryonic and blastoid potential

ABSTRACT: Naive human pluripotent stem cells (hPSCs) represent a pre-implantation epiblast state able to efficiently differentiate into embryonic and extraembryonic pre-implantation lineages and to self-organise into blastocyst-like structures called blastoids. Naive hPSCs maintenance commonly relies on mouse embryonic fibroblast (MEFs), introducing variability and analytical confounders. Here, we describe a feeder-free culture system based on serum coating that supports long-term naive hPSC maintenance. Across five laboratories, 30 serum batches were evaluated for the expansion of 8 naive hPSC lines for up to 25 passages. Mass spectrometry identified fibronectin and collagens as extracellular matrix proteins consistently present in serum coating. Cells cultured on serum coating displayed growth kinetics, clonogenic capacity, mutation rates, and global gene expression profiles comparable to MEF-based cultures. Importantly, serum-cultured naive hPSCs efficiently underwent germ layer specification, retained trophectoderm competence, and generated blastoids with similar efficiency. Collectively, serum coating provides a scalable, cost-effective, and robust alternative to feeder-based systems, preserving genomic stability and developmental potential while eliminating MEF-associated disadvantages and variability. This platform facilitates large-scale applications of naive hPSCs and enables more reproducible mechanistic studies.

INSTRUMENT(S):

ORGANISM(S): Rattus Norvegicus (rat) Bos Taurus (bovine)

TISSUE(S): Early Embryonic Cell

SUBMITTER: Graziano Martello  

LAB HEAD: Graziano Martello

PROVIDER: PXD059820 | Pride | 2025-12-22

REPOSITORIES: Pride