General Description

BD FACSAria II is a high-speed fixed alignment benchtop cell sorter. It uses low-powered, air-cooled and solid-state lasers: Violet (407-nm, 30 mW), Blue (488-nm, 13 mW) and Red (633-nm, 11 mW). System is equipped with Forward (FSC) and side (SSC) scatter detectors and 9 fluorescence detectors (link). The tube holders for 1 ml, 1.5 ml, 5 ml and 15 ml are provided. BD FACSAria II can sort up to four populations at once. There is no multi-well plate sorting module. The variety of particle sizes can be accommodated thanks to integrated nozzles available in four sizes (70 µm, 85 µm, 100 µm & 130 µm). BD FACSAria II can analyze up to 70 000 events/sec.

Sample Preparation

Prepare cells suspension in filtered buffer (PBS with 2% BSA + 2.5 mM EDTA) but if necessary in any other medium without phenol red and FCS. The sample should have particles density of approximately 5-10 million/ml. If less cells are available put then into minimal volume of 500 µl. Negative controls and individual positive controls of your staining are required to set sorting gates appropriately.

Tips:

To improve cells viability make all preparation steps on ice, unless otherwise stated in your protocol.
To exclude dead cells from the analysis or sorting it is recommended to use DNA-binding dyes as a marker of dead cells.
To reduce cells aggregation in your samples incubate the cells in the presence of DNAse I (100 ug/ml with 5 mM MgCl₂). DNA released from dead cells during cells dissociation cause stickiness.

General Description

Sample Preparation

Further Reading