Complexity of lung microenvironment and changes in cellular composition during disease make it exceptionally hard to understand molecular mechanisms driving development of chronic lung diseases. Although recent advances in cell type–resolved approaches hold great promise for studying complex diseases, their implementation relies on local access to fresh tissue, as traditional tissue storage methods do not allow viable cell isolation. To overcome these hurdles, we developed a versatile workflow that allows storage of lung tissue with high viability, permits thorough sample quality check before cell isolation, and befits sequencing-based profiling. We demonstrate that cryopreservation enables isolation of multiple cell types from both healthy and diseased lungs. Basal cells from cryopreserved airways retain their differentiation ability, indicating that cellular identity is not altered by cryopreservation. Importantly, using RNA sequencing and EPIC Array, we show that gene expression and DNA methylation signatures are preserved upon cryopreservation, emphasizing the suitability of our workflow for omics profiling of lung cells. Moreover, we obtained high-quality single-cell RNA-sequencing data of cells from cryopreserved human lungs, demonstrating that cryopreservation empowers single-cell approaches. Overall, thanks to its simplicity, our workflow is well suited for prospective tissue collection by academic collaborators and biobanks, opening worldwide access to viable human tissue.


Maria Llamazares-Prada, Elisa Espinet, Vedrana Mijošek, Uwe Schwartz, Pavlo Lutsik, Raluca Tamas, Mandy Richter, Annika Behrendt, Stephanie T. Pohl, Naja P. Benz, Thomas Muley, Arne Warth, Claus Peter Heußel, Hauke Winter, Jonathan J. M. Landry, Felix J.F. Herth, Tinne C.J. Mertens, Harry Karmouty-Quintana, Ina Koch, Vladimir Benes, Jan O. Korbel, Sebastian M. Waszak, Andreas Trumpp, David M. Wyatt, Heiko F. Stahl, Christoph Plass, Renata Z. Jurkowska


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