Running the flow cytometer BD Accuri C6
- Choose between the number of events counted or velocity the cell culture is drawn through the flow cytometer
- Threshold – indicates which cells will be excluded from a count
- Wash cycles (miliQ water) – washing the nozzle
- Agitation cycles – agitation of the cell samples
- Fluorescence filters – 4 different FL1 533 ± 15 nm (488 ex.), FL2 585 ± 20 nm (488 ex.), FL3 670nm (488 ex.) and FL4 675 ± 12.5 nm (650 ex.)
- Manual setups – for one sample
- Automatic setups – for several parallel samples (24/96)
- 0.5 mL of sample is enough
- Log scales or linear scales
Cleaning steps
After cleaning steps the plots that supposed to appear on the screen can be compared with plots on pages 12-13 of User Guide book next to the flow cytometer. Add following solutions:
A1 – clean solution
A2 – decontamination solution
A3 – miliQ water
A4 – 8 beads solution (if you are making new one, take 8 drops from black flask + 1 ml miliQ water)
A5 – miliQ water
A5 – 6 beads solution (if you are making new one, take 4 drops from each of two white flasks + 1ml miliQ water)
(Beads are for calibration of the flow cytometer)
B1 – miliQ
Run “Template Wash and Beads Randi”
Settings:
MiliQ water A1, A2, A3, A5, B1 – 5 min, 1 cycle wash, 1 cycle agitation
A4, A6 – 10000 events, Slow, 1 cycle wash, 1 cycle agitation
Result analysis
- Forward scatter FSC – size of the cells
- Side scatter SSC – shape of the cells
- Only histogram plot shows cell count
- Dot plot – can show different populations according to their size, shape or florescence compound content