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Section of Immunobiology, Department of Internal Medicine and Yale Cancer Center |
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Cell Sorter Facility Policy
1. No primary human materials will be accepted for sorting unless documented proof (such as a test slip) of patient's negative results of testing for HIV, Hepatitis B and Hepatitis C will be provided prior to sort. This must be clear before time is reserved. FACSVantage SE, FACS Aria and FACS Aria Amistad are equipped with aerosol evacuation systems and therefore are suitable for handling human cells.
2. To book your sorting experiment, please call (203) 737-2891 for MoFlo, (203) 785-2851 for FACSVantage SE, (203) 785-2541 for FACSAria, (203)785-2299 for Aria at Amistad or stop by at Room S617 or Amistad 416. Before writing your name in the sorting schedule book, you must have a brief discussion with cell sorter operators about your experimental design (e.g. the fluorochromes being used, diagrams of the cell populations, amount of sorted material expected, and sorting regions desired). If you do not, your appointment may be canceled without notice. This is required to optimize the usage and set-up of the cell sorters.
3. Samples must be filtered through (40 u mesh) before sorting . For example, you can use the following products: B-CMN-40 (by sq. feet) from Small Parts, Inc.; CellTrics sterile filter caps from Partec, Falcon 2235 tubes with the mesh in the caps from Becton Dickinson, etc. The optimal concentration of material in the sample tubes: (15-17) x106 cells /ml. Cells bigger than lymphocytes might need to be at lower concentrations. The following measures are important to prevent sorting material from clumping: a) concentration of fetal calf serum or BSA should not exceed 0.5%; b) use filtered buffer that is Ca2+ and Mg2+ free unless measurement depends on these ions; c) add some 0.02% EDTA; d) use anti-clumping agents, such as Accumax. It is highly recommended that unstained and single color stained controls be provided for optimum set-up. Sufficient amount of collecting tubes (of 5 -ml or 15-ml capacity,) with media (up to 1ml or 3 ml per tube respectively) is necessary. To sort populations of low percentage, higher volume of media (up to 3 ml in 5-ml tubes) is recommended for better cell recovery. Media in collection tubes should contain at least 10% serum to increase viability of the cells. If many cells are expected to be collected, the serum concentration can be adjusted to100% for dilution with the sheath fluid.
4. For MoFlo sorts, cells should be brought in 12 x 75 mm polypropylene test tubes (available upon request at S617, BD cat.#352063). For collecting, any type can be used (5, 15, 50 ml test tubes and small Petri dishes). 5. In case of delay or cancellation, notify cell sorter facility personnel as soon as possible by phone or e-mail : Thomas.Taylor@yale.edu, Gouzel.Tokmoulina@yale.edu, Geoffrey.Lyon@yale.edu, Zhao.Zhao@yale.edu. Users are not allowed to erase their names by themselves without telling the staff. If your Monday sort is going to be cancelled, and you didn't know until the weekend, the best way to notify us is to leave a note on the Cell Sorter Room door, TAC S 617 or Amistad 416. Individuals who cancel sorts with <24 hrs notice on two occasions will not be able to reserve or use the machine for two months and if they repeat after this, they will lose the ability to reserve or use the sorters. 6. In order to assure more equitable access, we are implementing the following until further notice:
1) Max time per sort: 5hr. 2) Only 1 5hr slot per week. 3) Max of 12 hr total signed up in advance per week.
7. Charging policy Sorting time will be charged based on the number of reserved hours or the number actually used, whichever is more.
Users who cancel the sort less than 24 hours prior to the scheduled time will be charged for their entire scheduled sort time . Users who cancel their sort time less than one week before their scheduled time but more than 24 hours in advance will be charged a flat $50 cancellation fee. This new fee will hopefully deter users or labs who sign-up for large blocks for time to ensure availability, only to cancel their time at the last minute.
Sorting no-shows ( 1hr.late without notice) will be charged at double the rate. Even in the case of accidental loss of material ( cells died etc.) the full scheduled sort time will be charged.
For FACS Vantage SE and MoFlo : $55/hr
For the FACSAria as follows: Sorting (with Geoff Lyon/Zhao Zhao setting up and supervising): $55/hr Sorting independently: $18/hr Sorting (with Geoff/Zhao helping setup but independent after that): $55 first hour and $28/hr thereafter Analysis (independent): $18/hr
Evening sorting 5-9 pm: $83/hr with minimum 2 hours per sort requirement
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