Flow Cytometry and High-Speed Cell Sorting Laboratory

The Flow Cytometry Laboratory at Auburn University provides operator-managed and assisted support for flow cytometry analyses and high-speed cell sorting. The facility has both analytical and cell sorting instrumentation available including both a Beckman Coulter analyzer and cell sorter.

The Flow Cytometry Laboratory offers flow cytometry analysis on a Beckman Coulter Analytical Flow Cytometer – a CytoFLEX LX flow cytometer – which is used for the qualitative and quantitative measurement of biological and physical properties of cells and other particles. These properties are measured when the cells pass through one or multiple laser beams in a single-file. The CytoFLEX LX flow cytometer can perform up to 21 color marker analyses simultaneously. The system has the maximum of 6 lasers installed (375 nm, 405 nm, 488 nm, 561nm, 638 nm, 808 nm) and can analyze cells in 21 unique fluorescent color channels. The CytoFLEX LX is also equipped with a 96-well plate-loader for automated sample analysis or can use sample flow tubes. Volumes range from ~50 ul up to 4.5 ml. Analysis can be performed at up to 25,000 cells/sec. Analysis of the data is performed on CytExpert Analytical Software.

CytoFLEX SRT Cell Sorter

The AU Flow Cytometry & High-Speed Cell Sorting Core Laboratory has recently installed a new state-of-the-art compact benchtop cell sorter that is housed within a BSL2-competant laminar flow biosafety cabinet.

The CytoFLEX SRT Cell Sorter employs ultrasensitive avalanche photodiode detectors (APDs) that can drive 4-way positive cell sorting as well as the ability to capture aborts. The sorter is configured with 4 lasers and 15 fluorescent color channels greatly expanding on our previous equipment. A simple and intuitive compensation workflow that also allows adjustment in real-time is provided by the APD technology. The sorter also has a robotic arm that can sort into 96 or 384-well plates, with or without index sorting, and employs automated sort setup and monitoring throughout each sort. The custom designed Baker SterilGARD Biosafety Cabinet is a Class II Type Biosafety Containment Cabinet designed specifically for the CytoFLEX SRT Cell Sorter. The biosafety cabinet relies on the flow of air to provide protection of personnel and containment by an intake air velocity of 100 feet per min (fpm) or more through the front access. All air exhausted from the hood is HEPA (high efficiency particulate air) filtered. The biosafety hood is certified for BSL2 level sorting and will sort at up to 40,000 events per sec processing fluorescent events using a fully digital system with 7 decades data display using CytExpert software.

LASER & Optical Filter Specifications

  • Violet Laser at 405 nm at 90 mW
    • Optical Channels – 405/10, 450/45, 525/40, 610/20, 660/10, 780/60
  • Blue Laser at 488 nm and 50 mW
    • Optical Channels – SSC – 488/8, 525/40, 690/50 + Forward Scatter (Cell Size)
  • Yellow Laser at 561 nm and 30 mW
    • Optical Channels –561/6, 610/20, 780/60, 585/42, 710/50, 675/30
  • Red Laser at 638 nm and 100 mW
    • Optical Channels –660/10, 712/25, 780/60

Also available are post-acquisition analysis software packages that provide listmode playback of all experiments in real-time with full analytical adjustment, cell cycle analysis, histogram subtraction and/or overlays of multiple histograms, and color-backgating of discrete cell populations. Secure long term data storage on hard drive, CD, and the local network are also provided. Data analysis is also available.

Use the Flow Cytometry facility description above for grant proposals.

Cell Sorting Applications:

  • Up to 16 color 18 Parameter Analysis – 4 lasers
  • Cell & Tumor Antigen Markers
  • High Speed 1-4 Way Cell Sorting
  • Autofluorescent Proteins & GFP
  • Antigen or Ligand Density
  • Apoptosis
  • DNA/RNA Content & Cell Cycle Analysis
  • Cytokine Receptors & Synthesis
  • Viability
  • Single Cell/Plate Sorting

Resources

Clients can purchase filter microscreens here (Item U-CMN-52).

Contact

Rie Watanabe
Manager and Operator
250A Greene Hall
Auburn, AL 36849
Lab Phone:334-844-2711
rzw0041@auburn.edu

Dr. Richard Curtis Bird
Laboratory Director
255 Greene Hall
Auburn, AL 36849
birdric@auburn.edu