This protocol outlines methods to obtain leukocytes, bone marrow mononuclear cells(BMMCs), and peripheral blood mononuclear cells (PBMCs) for use with 10x Genomics Single Cell RNA Sequencing protocols. The protocol was demonstrated using whole blood and bone marrow aspirate collected in various BD Vacutainer tubes containing anticoagulants. Comparable postisolation cell viability was observed between various collection tubes with a range of anticoagulants. For best granulocyte quality and viability, whole blood should be processed within 2 h of collection.
Important
This demonstrated protocol is optimized for counting cells in the range of 700-1200 cells/µl. If using the Single Cell 3' LT v3.1 (low throughput) application, ensure cells are counted as indicated in this protocol and then diluted to the LT specific optimal loading concentration of 100-600 cells/µl using the Cell Dilution Overview in the LT User Guide.