Topic
Pseudoreplication
Are my cells (or spots, or technical replicates) really independent samples?
Treating 5,000 cells from 2 donors as 5,000 replicates produces p-values of 1e-200 for effects that would not replicate in a third donor. The unit of replication is the biological sample; pseudobulk aggregation or mixed models restore it.
- How to Avoid Pseudoreplication in Single-Cell ATAC-seq
Five thousand cells from two donors are not five thousand replicates, and your p-values are lying to you about it.
- How to Avoid Pseudoreplication in Single-Cell RNA-seq
A p-value of 1e-200 from FindMarkers usually means you tested cells instead of donors, and pseudobulk is the fix a reviewer already knows to ask for.
- How to Avoid Pseudoreplication in Single-Nucleus RNA-seq
5,000 nuclei from two donors is not 5,000 replicates: pick pseudobulk or a mixed model before you trust a single p-value out of FindMarkers.
- How to Avoid Pseudoreplication in Spatial Transcriptomics
A wall of p-values near zero on pooled spots or cells almost always means you counted the wrong thing as your sample size.