Optimal strategy to process samples to compare two different condition.

[singcell]

I wanted to compare 3 PBMC and 3 Stimulated samples of scRNA-Seq run. I was wondering what would be the best way to process the data:

1) Process each of 3 PBMC separately using CellRanger count and combine 3 control 10X Runs (Cellranger count outputs) using Seurat merge function to make PBMC Seurat object and do same with Stimulated samples. Then compare control and treated samples using Stimulated vs Control PBMCs vignette .

Or

2) Use CellRanger aggr to aggregate 3 control and 3 treated 10x runs in two separate PBMC and Stimulated CellRanger aggr outputs and make two different Seurat object and then use Stimulated vs Control PBMCs vignette.

[timoast]

I wouldn't recommend using cellranger aggr as it will downsample everything to a similar number of counts as the sample with the lowest counts, and so potentially will throw out a lot of data. You could quantify each dataset (using cellranger, or another tool like Alevin) and first merge them in Seurat and check if there are batch differences between the datasets. If there are, you could run the integration methods