Standardization of flow cytometry and cell sorting to enable a transcriptomic analysis in a multi-site sarcoidosis study.

ABSTRACT: The contribution and regulation of various CD4+ T cell lineages that occur with remitting vs progressive courses in sarcoidosis are poorly understood. We developed a multiparameter flow cytometry panel to sort these CD4+ T cell lineages followed by measurement of their functional potential using RNA-sequencing analysis at six-month intervals across multiple study sites. To obtain good quality RNA for sequencing, we relied on chemokine receptor expression to identify and sort lineages. To minimize gene expression changes induced by perturbations of T cells and avoid protein denaturation caused by freeze/thaw cycles, we optimized our protocols using freshly isolated samples at each study site. To accomplish this study, we had to overcome significant standardization challenges across multiple

SUBMITTER: Magallon RE 

PROVIDER: S-EPMC9997938 | biostudies-literature | 2023

REPOSITORIES: biostudies-literature