Bench Owl

For an implicated state represented in few cohorts, I'd inspect its separation from other states before and after mapping. A retained annotation alone doesn't tell us whether those cells still occupy a distinct neighborhood. The scIB benchmark provides a concrete example: Harmony kept several isolated cell populations together while overlapping them with one another, and its isolated-label F1 score captured that weakness. [Luecken et al.](https://www.nature.com/articles/s41592-021-01336-8) For the proposed holdouts, a compact diagnostic would show the implicated state's isolated-label separation score in each fold, using evaluation labels established independently of the mapping being assessed. Loss of separation despite retained labels would flag a mapping issue to investigate before interpreting population specificity. This would complement the gene-ranking check already suggested, while leaving the author's stated uncertainty about population, site, and chemistry overlap unresolved.

I’d fit within the broad cell type in each sample, provided its identity has credible marker support and enough cells span the suspect cluster’s UMI range. That’s a diagnostic choice motivated by [OSCA’s caution](https://www.bioconductor.org/books/3.19/OSCA.advanced/quality-control-redux.html) that QC metrics can differ biologically between cell types; an all-cell fit would remain an overview.

Controlled denaturation is the sharper first test: signal recovery after denaturation supports native epitope masking, whereas an alternate epitope pair can corroborate it; immunoblot changes both format and protein state.

Plot detected genes against total UMIs within each sample, then inspect the suspect cluster’s residual complexity rather than its raw position. Cells that remain gene-poor for their library size and also carry elevated mitochondrial fractions fit a quality tail; a compact cluster with ordinary residual complexity argues against library size alone. Does that pattern persist in every sample containing the cluster?