BD LSRII Maintenance Guidelines

  1. Login into Windows using your RUNet Login and password.
  2. Check if the on-screen user interface shows BD LSRII and FACSFlow system "ON". If it is "OFF" be sure to click only upper button once to switch "ON" both BD LSRII and FACSFlow system.
  3. Please make sure that the green lights on both - instrument BD LSRII and FACSFlow system are actually "ON".
  4. If the fluorochromes/dyes you use in the experiment require the UV or 445 nm lasersto be "on" please make sure to follow the appropriate steps:
    1. If you use the UV-excited fluorochromes/dyes (Indo, Hoechst, AF350, CellTrace Calcein Blue AM, LIVE/DEAD Fixable Blue, etc.) please make sure that the UV laser on the BD LSRII-1 is "on" by following the steps in "The Operation Procedure for UV Laser on BD LSRII-1";
    2. If you use the 445nm-excited fluorochromes/dyes (CFP, mCerulean, etc.) please make sure that the 445 nm laser on the BD LSRII-2 is "on" by following the steps in "The Operation Procedure for 445 nm Laser on BD LSRII-2".
  5. Before you start running samples please:
    1. Check if the BD LSRII fluidic system is functioning properly:
      1. Remove the tube with MQ water from the sip (Sample Injection Port);
      2. Push "RUN" and "High" fluidic control buttons;
      3. Check if the buffer starts dripping from the sip:
        • If "yes" - please proceed to the step 5b;
        • If "not" - please push the "Standby" button immediately and report the problem to FCRC Staff (see the notes on the end of the document).
    2. Check if the trap filter attached to the pressurized plastic tank is free of air bubblesby bleeding sheath fluid from two ports:
      1. Open the roller clamp on the top of the trap/bubble filter for 3-5 seconds;
      2. Sequentially open the second roller clamp (at the right side of the instrument) for 3-5 seconds;
      3. Check if you are getting consistent stream release from both ports:
        • If "yes" - please proceed to the step 5c;
        • If "not" - you should suspect pressure problems or presence of bubbles, please immediately push the "Standby" button and report the problem to FCRC Staff (see the notes on the end of the document).
    3. Wash the instrumentbefore you run samples:
      1. "Run" at "High" 2 ml of 10% Bleach for 5 minutes;
      2. "Run" at "High" 2 ml of MQ Water for 5 minutes;
      3. Push "Standby" button and leave tube with 1 ml of MQ Water on the sip.
        Note: Please be sure in all the cases not to exceed the maximum allowed volume of 2 ml in the sample tube.
  6. Login into DiVa. Always choose to use CS&T settings. Create new experiment, or import the old one and duplicate without data. Run your experiment. Export data. Copy them to the CFS. Delete your experiment from the DiVa Database. Logoff DiVa to avoid unnecessary charges.
  7. When you are done with the experiment if you have used the UV or 445 nm laser be sure to switch it off. Refer to the step 3 in "The Operation Procedure for UV Laser on BD LSRII-1" or "The Operation Procedure for 445 nm Laser on BD LSRII-2", respectively.
  8. When you are done with the experiment be sure to wash BD LSRII properly:
    1. "Run" at "High" 2 ml of 10% Bleach for 10 minutes;
    2. "Run" at "High" 2 ml of MQ Water for 10 minutes;
    3. Push "Standby" button and leave tube with 1 ml of MQ Water on the sip.
  9. Before you leave FCRC please be sure to switch instrument off if required. Policy of keeping instruments "ON" or "OFF" depends on the time of the day:
    1. During the daytime, we keep the analyzers "ON" all the time;
    2. If you finish your BD LSRII experiment after 6 pm on the weekday or any time on the weekend and if the next person is not physically present at the site we request to switch "OFF" the BD LSRII and FACSFlow system by using the on-screen user interface. Please click only upper button once to switch "OFF" both BD LSRII and FACSFlow system.
  10. Please be sure to logoff Windows before you leave FCRC. Otherwise in 10 minutes Windows will automatically lock the session under your name and will prevent other researchers from usage of the instrument.

Note: Please report all the problems/concerns to FCRC Staff:

During business hours — get to DWB205 and tell the FCRC staff member.
After hours — e-mail to fcrc@rockefeller.edu; leave the note about the problem on the instrument's keyboard; proceed on step 9b from the above list.