The challenges of single cell transcriptomics on difficult human tissue: the placenta
Xenakis, T.; Moreno Villena, J. J.; Hall, G. T.; Hillman, S. L.; Sanchez Corrales, Y. E.; Castellano, S.
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Demand for the application of single cell transcriptomics on difficult tissues, processed and stored in disparate conditions, has led to the development of various single cell modalities. We focus on the placenta, a challenging tissue to transcriptomically interrogate due to senescence, intermittent hypoxia, high levels of RNase activity and tissue trauma at delivery. We performed single cell and nuclei transcriptomics on two samples with probe-based and native molecule transcript capture, droplet based and in situ plate based cell isolation. We explored sample and storage variations, including freshly dissociated tissues, fixed cells, snap frozen and FFPE processing. We find that variations in sample processing and storage have much larger effect on cell type proportions than differences in chemistry, impacting the biology that can be learned or the identification of disease markers. Further, the transcriptomic output of in situ combinatorial indexing consistently overlaps, with single nucleus transcriptomics, sharing little with other modalities. This may limit combinatorial indexing for sampling cytoplasmstic mRNAs. Our comprehensive analysis of the varying effect of single cell transcriptomic modalities, including their sample management strategies, provides novel and essential considerations for the experimental design and analysis of single cell transcriptomics applicable to challenging tissues, offering actionable guidance for future experiments. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=76 SRC="FIGDIR/small/688483v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@a0767forg.highwire.dtl.DTLVardef@c3af0corg.highwire.dtl.DTLVardef@1193e3eorg.highwire.dtl.DTLVardef@45989c_HPS_FORMAT_FIGEXP M_FIG C_FIG
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