Question: What buffers can be used for washing and cell resuspension?
Answer: To prepare single cells for Chromium Single Cell applications, it is recommended to use 1X PBS (calcium and magnesium free) containing 0.04% weight/volume BSA (400 µg/ml) for washing and resuspension. Fresh and frozen/thawed PBMC samples as well as cell lines have been tested with this buffer.
Primary cells, stem cells, and other sensitive cell types may require washing and resuspension in alternative buffers to maximize viability. For other buffers that have been tested, please refer to the Cell Preparation Guide (https://support.10xgenomics.com/single-cell-gene-expression/sample-prep/doc/demonstrated-protocol-single-cell-protocols-cell-preparation-guide). It is also possible to use most cell culture media with up to 10% FBS or up to 2% BSA to maintain cell health with little to no adverse downstream effects.
Media should not contain excessive amounts of EDTA (> 0.1mM) or magnesium (> 3mM) as those components will inhibit the reverse transcription reaction.
Any surfactants (Tween-20, etc) should also be avoided as they may interfere with GEM generation.
When using untested buffers or cell culture media, it is recommended to minimize the volume of cell suspension volume added to the Single Cell Master Mix. This will reduce carryover and potential adverse effects of any buffer components.
Products: Single Cell 3', VDJ