Author
Date
Version
SOP #
Stephanie Swift
January 2014
1
G003
Objective: This SOP addresses the maintenance and upkeep of the FACSCanto flow
cytometer
SOP-G003: FACSCanto Maintenance Schedule and Frequency
A
B
C
D
E
Maintenance
System Initialisation
Fluidics Cleaning
Empty Waste
Laser
System Flush
Frequency
Before Use
After Use
When Full
Weekly
Bi-Monthly
Note: to improve cytometer performance and minimise clumped cells in your
sample population (which will falsely skew your data), all samples should be
filtered through 48um nylon mesh before running on the flow cytometer.
A. SYSTEM INITIALISATION
1. Turn on FACSCanto flow cytometer using the green button on the front of
the instrument.
2. Open Diva software and check to ensure that the FACSCanto flow
cytometer is connected to the computer.
3. Perform fluidics start up (approx. 10 mins.)
4. Wait at least 30 minutes for the lasers to warm up and stabilise before
running samples.
5. Perform Cytometer Setup and Tracking (CST) (should be performed by an
admin user at the beginning of the day, before running any experiments).
Invert the CS&T bead tube several times to mix and add 1 drop of CS&T
beads to 400ul of sheath fluid in a 5ml polystyrene tube. In DIVA, select
CS&T, confirm that the bead lot number is the same as the lot selected on the
screen. Place the CS&T bead mixture on the SIP and select run on the
cytometer. Upon completion select use CST settings.
Note: Each new lot of CS&T beads has to have a baseline tracking set-up
before they can be used. This, like the CS&T itself, can only be done by an
admin user.
This protocol is shared by the Stojdl Lab under a creative commons license.
B. FLUIDICS CLEANING
1. At the end of each use, wipe the SIP clean.
2. Install and run a tube containing 3 mL of a 1:10 dilution of bleach on the
SIP. Allow the sample to aspirate for approximately five (5) minutes.
3. Remove tube, then install a second tube containing 3 mL dH2O on the SIP.
Allow the vacuum to aspirate approximately five (5) minutes.
4. Repeat step 3 with a third tube containing 3 mL dH2O.
5. Place the instrument on STNDBY and continue with Shutdown if needed
(i.e. if nobody is running less than 2 hours after you).
C. EMPTY WASTE
Empty waste containers as required (the machine will prompt you when
emptying is necessary), and make sure 10% bleach is added into the waste
container before it gets replaced on the wet cart stand (enough to cover the
bottom of the container).
a) Place instrument on STANDBY before disconnecting waste.
b) Disconnect orange pressure tubing by pressing on the metal tab.
c) Remove the cap at the sink in the room and pour the waste down
the drain while running water to flush the drain.
d) Add fresh bleach to cover the bottom of the container.
e) Re-connect the tubing when finished.
f) Use appropriate lifting technique for waste container.
D. LASER MAINTENANCE
To extend the life of the lasers, turn on the instrument for 3 – 4 hours at least
once per week.
E. SYSTEM FLUSH PROCESS
An overall fluidics cleaning is required to remove debris and contaminants
from the sheath tubing, waste tubing, and flow cell. Perform the system flush
at least once every two (2) weeks.
Note: Do not run detergent, bleach, or ethanol through sheath filter, this may
break down the filter paper within the filter body, causing particles to escape
into the sheath fluid, possibly clogging the flow cell.
1. Remove sheath filter (bypass filter).
2. Empty sheath container and rinse with deionized water.
This protocol is shared by the Stojdl Lab under a creative commons license.
3. Fill sheath container with at least 1 liter of a 1:10 dilution of bleach or fullstrength BD FACSClean™ solution.
4. Empty the waste container it is ≥ ¾ full.
5. Install a tube with 3 mL of 1:10 bleach or full-strength BD FACSClean™
solution and acquire data on high for 30 minutes.
6. Remove tube and depressurise the sheath container by lifting the pressure
relief valve cap.
7. Empty sheath container and rinse with deionized water.
8. Fill sheath container with BD FACSRinse™ solution.
9. Install a tube with 3 mL of full-strength BD FACSRinse™ solution on the
SIP.
10. Repeat steps 5 – 7 with BD FACSRinse™ solution.
12. Fill sheath container with at least 1 liter of deionized water.
13. Repeat steps 5 – 7 with deionized water.
14. Replace sheath filter and refill sheath container with sheath fluid.
Admin user contacts:
1. Katelynn Rowe, Stojdl Lab (RI2, fourth floor)
katelynn@arc.cheo.ca
2. Stephanie Swift, Stojdl Lab (RI2, fourth floor)
stephanies@arc.cheo.ca
This protocol is shared by the Stojdl Lab under a creative commons license.
FACSCanto Flow Cytometer Maintenance Log
Check When Complete
Date Cleaned
(dd/mm/yyyy)
Initials
& Lab of
Cleaner
System
Initialisation
Fluidics
Cleaning
Empty
Waste
Laser
System
Flush
This protocol is shared by the Stojdl Lab under a creative commons license.
Notes