Question: Why is the recovery agent not completely removed after breaking emulsions?
Answer: After removing the Recovery Agent/Partitioning Oil (pink layer) from the bottom of the sample and discarded, 10 to 15 µl should remain:
The Single Cell 3' protocol was designed to have 10 to 15 µl of Partitioning Oil left so that there is no risk of removing any of the aqueous layer. Removal of the aqueous phase will result in a significant decrease in library complexity.
Note: Remainder volumes >15 µl can indicate a clogging failure.
Products: Single Cell 3', VDJ