University of Vermont

College of Medicine

Flow Cytometry and Cell Sorting (FCCS) Facility

Sorter

Sorter

  • Please, read Practical Cell Sorting before doing a sort!
  • Sample tubes: The following tubes may be use to sort: 1 ml microtubes (BioRad, cat # 223-9391), 5 ml/12x75mm polystyrene round bottom tubes (Corning non-sterile cat# 352008 or 352052; sterile option cat# 352054 or 352003), or 15 ml centrifuge tubes (Corning: polypropylene cat # 430052, 430766, 430790; polystyrene: cat # 352099).
  • We recommend bringing your samples in a buffer containing ≤ 1% of BSA, FBS, etc.
  • Sort collection tubes: provide 5 or 15 ml protein-coated polypropylene tubes. To coat tubes, fill up tubes with serum containing medium and let sit ≥ 1 hour prior to the sort. Discard all but aprox. 1 or 2 ml of media. If cells are especially sensitive to serum concentration we recommend using 2x the amount of serum. Alternatively, collection tubes can be coated with 100-200 µl of FBS (be sure FBS covers/touches the entire inner wall of the tube). Cells can also be sorted directly into 384-96-48-24 well plates.
  • Filter/cell strainer: Samples on the cell sorter MUST be filtered before running them (even without visible aggregates). The nozzles are easily clogged. 35 to 70 µm nylon mesh or cell strainer are mandatory (CellTreat, Corning cat # 352235, nylon mesh from Partec cat # 04-004-2327, and Smallparts). We reserve the right to charge for removing clumps from the machine in addition to the lost machine time.
  • Gating strategy: The user is required to discuss the gating strategy of the sorting with the FCCS staff prior to the sort. It is the responsibility of the investigator to specify the cell population to be sorted.
  • Sample: You must bring a negative control, as well as individual positive controls for each fluorochrome you are using. The FCCS facility does not provide controls for your experiment. Please, refer to our FC Protocols > Compensation link.
  • Forms: All users sorting samples on the FACSAria must fill out and return the Sort Request Form and FCCS Biohazard Sort Form 48 hours before your scheduled sorting.
  • BSL2 samples: For all BSL-2 material, users must adhere to proper “Transportation of potentially infectious material environmental safety protocol” (UVM-Safety) and must dispose of all biowaste in their own lab. Samples must be contained in a leak-proof container and labeled with a biohazard sticker. Samples should be transported on campus in a covered, sealable, secondary container (e.g., Tupperware or screw top, gasketed plastic jar). Gasketed jars can be obtained by contacting Environmental Safety. In addition to secondary containment, tertiary containment - a hard plastic, locking cooler (for example, an Igloo cooler) - is required for transportation of infectious or potentially infectious material outside of the laboratory. Contact us if you need a locking cooler to transport BSL2 samples to this facility.

Last modified August 06 2015 03:08 PM