dead space the marker dead space marker language

Notably, both single-cell and single-nuclei RNA-seq do not address the gap between RNA and protein expression, which can result from technical capture difficulties (lack in transcript capture/drop-out) and biological processes (translational impediments, post-translational influences, RNA degradation kinetics or protein trafficking to and from the cell surface) 7, 8. Single-nuclei RNA-sequencing has been proposed for larger cohorts of fresh-frozen samples or where tissue dissociation is hard to achieve, such as for mature interconnected brain tissues 3, 4, 5 but this is not without its drawbacks 6. As an unbiased technology that does not profile a panel of preselected transcripts, it is ideally suited to resolve cellular heterogeneity in complex healthy or diseased tissues. The last decade has been marked by immense progress in the field of single-cell RNA sequencing (scRNA-seq) 1, 2. This work provides practical guidelines for processing and analysis of clinically relevant solid tissue biopsies for biomarker discovery. Finally, in a proof-of-concept study, using protein abundance analysis on selected surface markers in a cohort of healthy skin, primary, and metastatic melanoma we identify CD56 surface marker expression on metastatic melanoma cells, which was further confirmed by multiplex immunohistochemistry. To further assess the RNA-protein gap, RNA-protein we perform codetection and correlation analyses on thresholded protein values. To assess the effect of enzymatic digestion on transcriptome and epitope expression in immune cell populations, we analyze peripheral blood mononuclear cells (PBMCs) with and without dissociation.

Next, we compare five skin dissociation protocols by taking into account dissociation efficiency, captured cell type heterogeneity and recovered surface proteome. Applying CITE-seq to clinically relevant solid biopsies to characterize healthy tissue and the tumor microenvironment is an essential next step in single-cell translational studies. In this study, gating of cell populations based on their transcriptome signatures for use in cell type-specific ridge plots allowed identification of positive antibody signals and setting of manual thresholds. Multi-omics profiling by CITE-seq bridges the RNA-protein gap in single-cell analysis but has been largely applied to liquid biopsies.

Dead space the marker dead space marker language