After the third round barcoding and before lysis, cells will be pooled and subsequently split into between one and eight distinct populations we call sublibraries. Each sublibrary will contain a different 4th barcode (the Illumina index) and can be sequenced separately. You will count cells before splitting them into your sublibraries, and thus you can choose the number of cells in each. In practice it can be useful to have at least one sublibrary with few cells; This can provide an estimate of gene detection per cell at deeper sequencing while using fewer sequencing reads.
