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Diagnostic Testing Lab
COMPARATIVE MEDICINE INSTITUTE

Flow Cytometry and Cell Sorting

 If you are looking for the Clinical Pathology & Immunology diagnostic testing lab service please click here.

Lab Contact:    919.513.6443  |     seschuet@ncsu.edu

The NCSU Flow Cytometry and Cell Sorting Laboratory is located on the third floor of the main College of Veterinary Medicine building. The laboratory provides instrumentation and assistance for multi-parameter flow cytometry analysis and sorting. It is available for use to all NC State researchers and to those outside the University community and operates as a by appointment, fee for service laboratory.

Services Available

The Flow Cytometry Facility offers the following services:

  • Experimental planning (consultation and panel design)
  • Instrument training (Cytoflex or LSRII)
  • User independent acquisition and analysis
  • Facility assisted acquisition and analysis
  • Facility performed cell sorting

RATES

  • Independent use on Cytoflex or LSRII: $55/hour
  • Facility assisted use on Cytoflex or LSRII: $150/hour
  • MoFlo sort setup fee: $35 (one time fee charged each sort regardless of length of sort)
  • MoFlo sort fee: $125/hour

There is a 30 minute minimum charge on the analyzers and a hour minimum charge for the sorter. After the minimum charge, experiments are charged in 15 minute increments. It is up to the individual to establish an account with the facility when becoming a new user; it is also the responsibility of the user to update any funding information.

Applications

Flow Cytometry is a technology used to analyze characteristics of particles as they pass by one or more lasers. Flow Cytometry can be used in a multitude of different applications. Listed below is a wide range of examples.

  • Apoptosis (quantification, measurement of DNA degradation, mitochondrial membrane potential, permeability changes, caspase activity)
  • Cell surface antigens
  • Characterizing multidrug resistance (MDR) in cancer cells
  • Chromosome analysis and sorting (library construction, chromosome paint)
  • Intracellular antigens
  • Oxidative burst
  • Protein expression and localization
  • Total DNA content (cell cycle analysis, cell kinetics, proliferation, ploidy, aneuploidy, endoreduplication)
  • Transgenic products in vivo, particularly Fluorescent Proteins

Instrumentation

ANALYZERS

Becton Dickinson LSRII: Has three lasers, 488 nm, 635 nm and 405 nm. Can analyze up to nine colors and utilizes BD Diva software for acquisition. Scheduling for the LSRII is available by online invitation calendar. Click here to access the LSRII configuration for experiment planning.

Beckman Coulter CytoFlex: Has a 488nm laser as well as a 633nm laser. It has the ability to analyze 3 parameters off the 488nm laser and one parameter off the 633nm laser. The CytoFlex is customizable for specific experiments. The Cytoflex can perform absolute counts. Scheduling is available by online calendar. Click here to access the available filters.

Excitation Fluorescence Channels Fluorochromes
488nm 525/40 BP Fitc, alexa Fluor 488, CFSE, Fluo-3
585/42 BP PE, PI
610/20 BP ECD, PE-Texas Red, PE-CF594, PI
690/50 BP PC5, PC5.5, PerCP, PerCP-cy5.5, PI
780/60 BP PC7
638nm 660/20 BP APC, Alexa Fluor 647, eFluor 660
712/25 BP APC-Alexa Fluor 700, Alexa Fluor 700
780/60 BP APC-Alexa Fluor 750, APC-Cy7, APC-H7, APC-eFluor 780
405nm 450/45 BP Pacific Blue, V450, eFluor 450, BV421
525/40 BP Krome Orange, AmCyan, V500, BV510
610/20 BP Violet610, BV605, Qdot 605
660/20 BP Violet660, BV650, Qdot 655
780/60 BP Violet780, BV785, Qdot 800

HIGH SPEED CELL SORTER

Beckman Coulter MoFlo XDP

The Beckman Coulter MoFlo XDP provides NCSU with the flexibility to perform a wide range of cell sorting experiments. Our XDP is configured with a 200mW 488nm laser, a 35mW 640nm laser, and two obis “plug and play” lasers; a 100mW 405nm laser and a 50mW 561nm laser. The sorter is capable of analyzing up to 10 colors plus forward and side scatter. In terms of sorting capability, the XDP can sort for up to four different cell populations at once and is capable of purifying populations that are less than 1% of the total input population. We have the ability to sort a wide variety of cell types due to the variable sizes of our interchangeable nozzle tips. Scheduling is only available through the flow cytometry facility manager. Click here to see the full capabilities of the instrument from Beckman Coulter.

 

SOFTWARE

The Flow Cytometry Facility offers multiple software options free of charge for users. Currently the facility has Cyt-Expert (PC based), FACSDiva (PC based), FCS express with multi-cycle analysis capabilities (PC based), Summit (PC based) and Kaluza (PC based). Cyt-Expert, Summit, and FCS express are available to users on their own PCs while Kaluza and FACSDiva are only available in the facility. Please contact the facility manager, Sarah Schuett, to discuss your software needs and determine which software will be best for your experiments.

POLICIES

  • Users must schedule appointments directly with the facility manager, Sarah Schuett via email, as far in advance as possible for cell sorting and facility assisted experiments.
  • Users who have been trained for independent operation of the analyzers will be given access to an online calendar system where they can schedule their appointments and will be given 24 hour access to the facility.
  • All appointments require 24 hour notice for cancellations. If you do not notify the facility manager in writing, you will be charged.
  • For first time users, it is required to make an appointment with the facility manager to discuss your proposed experiment to be sure the laboratory has the capabilities to acquire the information you wish.
  • The Flow Cytometry Facility is a BSL-1 facility. Human (this includes established human cell lines as well as primary cells) and non human primate samples may be run as long as they are fixed. If you have questions about this, please contact the facility manager or EHS.
  • Bills are broken down by specific user and instrument use per date and are sent out monthly.
  • Users of the facility will automatically be added to a flow cytometry listserv that will keep users up to date with the state of the facility.

Google Calendar here

Sarah Bianco

Population Health and Pathobiology

seschuet@ncsu.edu

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