University of Vermont

College of Medicine

Flow Cytometry and Cell Sorting (FCCS) Facility

Analyzers

General considerations

  • Clogging of the flow cytometer fluidics is a serious problem that not only jeopardizes your own experiment, but can jeopardize the experiments from multiple laboratories that need to use the instrument that day. Proper sample preparation will minimize the chances of instrument clogs.
  • Filter/cell strainer: If there are clumps you can see in the sample tube, or if your cells were treated with trypsin, then you will need to filter your samples because they are likely to clog the instrument. Nylon mesh or cell strainer are highly recommended (≤70 µm; CellTreat, Corning cat # 35-2235, nylon-mesh filters from Partec (CellTrics), Smallparts, and Porex Brand SQ-Easy PreFilter System Columns).
  • Gloves MUST be worn when using the cytometer.
  • You must bring your own material, such as tubes, pipettes and tips, we do not provide them.

BD LSRII

  • Tubes: The LSRII requires Falcon 12x75mm polystyrene round bottom tubes (Corning non-sterile cat# 352008 or 352052; sterile option cat# 352054 or 352003). Small tubes designed to fit in a 96-well format holder (USA Scientific cat # 1773-2022) can also be used. Other options may be discussed.
  • Cell Concentration: For immunophenotyping analysis, apoptosis, or DNA content the final concentration should be 1-3 x 10^6 cells/ml. The minimum volume that can be run using a 12x75 mm tubes is 300 µL and 40 µL using small tubes (1 ml).
  • Preparation of Samples: Refer to General Surface Staining and Compensation Controls in our Flow Cytometer Protocols link.

MACSQuant VYB

  • Tubes: It can be used any 12x75 mm (5 ml) round bottom tubes and 1.5 ml centrifuge tubes.
  • Plates: 96 U-bottom and flat-bottom well plates
  • Cell Concentration: For immunophenotyping analysis, apoptosis, or DNA content the final concentration should be 1-3 x 10^6 cells/ml; however it can work with less. The minimum optimal volume to acquire is 25 µL off 50 µL.
  • Cell count: This instrument has the option to provide absolute cell count events/µL or events/mL.

Last modified August 06 2015 11:11 AM