Rules for Flow Cytometry Analyzers

The policies described below are put in place to delivery the lowest priced services possible based on cost recovery. Machine "hoarding" (scheduling a large amount of time with no intention of using all of the scheduled time) and then cancelling all or a portion of the time, is not fair to other researchers. This has been an issue in the past and slowed the pace of research on campus by preventing others from having access to machines. If this were this to go unchecked, it would result in higher fees for all investigators.

1. Users must quit the login tracking software, AppGate, after each session to avoid being charged for the time that someone else is using.
2. If the next user arrives at their scheduled time you must relinquish the machine to the scheduled user.
3. A single person cannot sign up for more than 8-hours per week of "prime time" (i.e. no more than 8 hours combined). Prime time is Monday-Friday 12N-6PM. An unlimited amount of time may be reserved out of "prime time". No more than 4 hours of "prime-time" can be reserved in any single block. If more than 4 hours of time is needed, individuals should sign up to start or end outside "prime-time". If your experiments require use of the analyzers beyond these limitations, please contact Diane Trotta. Exceptions will be determined on a case-by-case basis.
4. Users will be charged for the greater of time scheduled or actual time used on the cytometers.
5. If a user is not going to be able to make a scheduled time, they must delete the event from yaleflowscheduling.com at least 3 hours before the scheduled time in order to avoid being charged for the time they had reserved for their use. In the event that another investigator uses all or a portion of a cancelled time, the person with the original reservation will only be responsible for the remaining time not utilized.
6. Facility staff reserves the right to suspend privileges of users who scheduled time with someone else's account information, as well as the individual(s) who lent their password.
7. Users cannot sign-up for time and give it to a fellow lab member, except in the case of a collaborative study and if facility staff is notified in advance (i.e. two individuals working legitimately together on a single experiment). The online scheduling system will report these events as No Shows and users will be billed as such without advance notice. Long term collaborative projects can be managed through other approaches. Please contact facility staff to make arrangements.

8. Users should always wear gloves when using the flow cytometry analyzers or their associated computers. Propidium Iodide and other potentially hazardous agents are regularly used at cytometers and the common use lab environment.
9. No eating, drinking, food or beverages are allowed in any facility room at any time.
10. Users analyzing bacteria, potentially infectious materials, or non-human primate, or human samples that have not been tested for common human pathogens such as HIV or HepB must fix these materials with an appropriate fixative (generally 2-5% paraformaldehyde) before analyzing them on any facility analyzer. If an individual is found to be running any of the above mentioned materials unfixed on a cytometer, their facility privileges may be suspended. Users who need to run the above materials unfixed must have them analyzed on a cell sorter with an aerosol evacuation system (except for yeast which cannot be run).

11. All samples run on the LSRII must be filtered at the machine just before running them, since the LSRII is sensitive to clogging. If users have a special consideration regarding this rule, please contacts the Flow Cytometry Facility Director, Dr. Ann Haberman. Anyone found running samples without filtering will be given a Level II violation. Repeat offenders can lose their facility privileges.
12. Filtering can be accomplished with Falcon Mesh Top 2235 tubes (Becton Dickinson). Please review our page for filtering protocols.
13. All users must run a short series of washes after finishing their experiment. This serves to flush out cell aggregates sticking to the fluidics, as well as residual fluorophores. This is ESPECIALLY important when using Propidium Iodide or Acridine Orange that are particularly sticky within the flow cell and must be inactivated by bleach to prevent carryover to the next user's samples.
    • 10% bleach for 5-minutes (1-min with with arm to the side, then 4-minutes with arm under the tube)
    • 5% Contrad soap for 3-minutes
    • Water for 3-minutes.
14. Users must empty the waste tank and add 100 ml of Clorox bleach to the waste tank after each use. Users must also refill the sheath fluid tank after each use. Users do not have to perform these tasks if:
    • The next user is waiting to use the machine and has agreed to perform these tasks after their use.
    • If the waste container is less than 1/3 full and the sheath is more than half-full. 15.
15. Do not leave the machines running or allow the sheath fluid to run dry. This causes air bubbles in the systems that are difficult to remove.
16. It is the user's responsibility to promptly export their own data. Exported data files will be kept in the FSC folder on the hard drive of the computers for one week. After one week all exported files will be deleted. Data in the DIVA database will be available for one month. After one month the database is purged to optimize the performance of DIVA. Experiments and experiment templates which contain settings, compensation and labels will be deleted from user accounts after six months. Any files on the computer desktop will be promptly deleted.
17. The flow cytometry analyzers (Caliburs, LSRIIs, Stratedigms) are to be turned off after 6 PM weekdays unless the next user is physically present. They are to be turned off on weekends after using unless the next user is physically present in the facility Lab. This is to ensure that the cytometers are not left on overnight and on weekends when not being used.
18. To report any problems with any of the flow cytometers, please first try to contact Diane directly or e-mail her at Diane.Trotta@yale.edu.

Level I Violations are:

1. Leaving the Machine in the "Run" position
2. Leaving an overfilled tube on the machine (more than 1-2 ml dH2O)
3. Altering the machines without authorization (i.e. removing SIP Sleeves or Loader Cylinder, changing the filters)
4. Any gross abuse of the machines

Level I Fines are:

• First Violation: $100 Fine
• Second Violation: $250 Fine
• Third Violation: $350 Fine and FACS Facility privileges suspended for 2 weeks

Level II Violations are:

1. Not emptying the waste container
2. Not filling the sheath fluid container
3. Not properly preparing samples - clogging the machine repeatedly
4. Not depressurizing the system after use
5. Violating any sign-up rules
6. Not filtering the samples before running them on LSRII analyzers
7. Leaving a cytometer on overnight

Level II Fines are:

• First Violation: warning to user and PI via E-mail
• Second Violation: $100 Fine
• Third Violation: $250 Fine
• Fourth Violation: $350 Fine