University of Pittsburgh Cancer Institute (UPCI)

CCSG Acknowledgement

Required CCSG Acknowledgement

The NCI requires that publications acknowledge the UPCI CCSG support, and they are tracking compliance. If a UPCI CCSG-supported Shared Resource provided data used in your publication, please include the following statement in the acknowledgment section of your publication(s):

"This project used the UPCI [insert name(s) of shared resource(s)] that [is/are] supported in part by award P30CA047904."

Shared Resource Directors: Please make sure to include this statement on all of your order forms, contracts, etc. as a reminder to your users to acknowledge the UPCI CCSG support.


Sorting

Cell Sorting or FACS (Fluorescence Activated Cell Sorting) can be used to isolate high purity fractions of cells in suspension. The cell sorter combines multi-parameter flow cytometry with the ability to sort individual cells into several distinct populations. Sterile, functional cells are collected and can be returned to culture, utilized in functional assays or further characterized through genomic, proteomic or similar analysis. Up to 4 populations, defined by Boolean gates, can be sorted simultaneously. Alternatively, a single population can be deposited directly into a variety of cell culture plate formats (e.g., 96-well plates). The number of cells/well can be specified for each well, facilitating cloning, single cell experiments or limiting dilution experiments, where graded numbers of cells are plated.

High speed sorting allows for the processing of large samples, as millions of cells can be sorted in a very short amount of time. Cells can be sorted to optimize purity or recovery. For information concerning sorting of potentially biohazardous samples, please see the BSL2 Safety Manual.

Special User Guidelines for the MoFlo High Speed Sorter

First time users or experienced users with a new project should contact Michael Meyer before setting up an appointment.

Beckman Coulter MoFlo Astrios

Beckman Coulter MoFlo Astrios High Speed Sorter

The Beckman Coulter MoFlo Astrios is a 5-laser, high-speed (10,000-20,000 cells/second) sorter capable of 6-way sorting, plate deposition, and single cell sorting. Biocontainment in a custom designed Class II Baker cabinet permits sorting of live human and retrovirus-transfected cells. The sorter can be configured to suit various excitation and/or emission needs.

View fluorochromes for the MoFlo Astrios

Special User Guidelines for the MoFlo Astrios High Speed Sorter

First time users or experienced users with a new project should contact Michael Meyer before setting up an appointment.

Sample Requirements

  • All samples must be in single cell suspension. Cells should be filtered through a <70 μm nylon mesh filter to prevent clogging the nozzle if the cell size is less than 70 μm.
  • Sample may contain 1-3% FCS, or <=0.2% BSA.
  • Cell concentration ideally should be at least 1x107 cells/mL and can be up to 5x107 cells/mL for non-adherent cells (low cell concentrations slow down sorting).
  • Samples can be in a variety of tubes; ask the staff if you are unsure whether or not your choice is appropriate.
  • For the purpose of compensation for multicolor analysis or sorting, you must bring an unstained control tube plus single color stained tubes for each fluorochrome (>5% positive). Ask us about using IgG capture beads (BD CompBeads, Cat # 552843) instead of cells for single stained controls.
  • No radioactive material. The facility does accept unfixed human or non-human primate lentivirus-transfected cells, but investigators must meet with and discuss these experiments with facility staff. Please make an appointment if you need to perform a biohazardous sort.
  • Collection container (user supplies one of following options):
    • Tubes: 12 x 75mm, 1.5ml, 15ml or 50ml.
    • Plates: n=6, 12, 24, 96 (include PCR plate), 384 well or custom size.
    • Trays/dishes or slides.
  • Fill collection container (~10% of volume) with desired collection medium.
  • If collecting cells for molecular work, please contact Michael Meyer to discuss collection conditions to optimize your recovery.