Flow Cytometry and Cell Sorting
The Flow Cytometry and Cell Sorting facility at North Carolina State University – College of Veterinary Medicine (NCSU-CVM) provides instrumentation and assistance with multi-parameter flow cytometry analysis and cell sorting. It is available for use to all NCSU researchers and to those outside of the university community. The facility is equipped with two state-of-the-art analyzers (BD LSRII and a Beckman Coulter CytoFLEX), a MoFlo-XDP high speed cell sorter, and an AutoMACS magnetic cell sorter. The facility was also recently upgraded to support cell sorting of Biosafety level-2/2+ (BSL2/BSL2+) samples.
The NCSU-CVM flow core facility offers multiple software options free of charge for current users. Currently the facility has FlowJo, Cyt-Expert, FACSDiva, FCS express with multi-cycle analysis capabilities, and Kaluza. Also, the facility provides experimental design services to the users.
Users have the option of getting fully trained to operate the instrumentation independently or to pay for a facility-assisted run.
High Speed Cell Sorter
Beckman Coulter MoFlo XDP
The Beckman Coulter MoFlo XDP provides NCSU and to those outside of the university community 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 simultaneously, and is capable of purifying populations that are less than 1% of the total input sample. We have the ability to sort a wide variety of cell types due to the variable sizes of our interchangeable nozzle tips. Please contact us to discuss about your experiment and schedule a sorting.
Becton Dickinson LSRII: Has three lasers, 405 nm (Violet), 488 nm (Blue) and 635 nm (Red). Can analyze up to nine colors and utilizes BD Diva software for acquisition. Click here to access the LSRII configuration for experiment planning.
Beckman Coulter CytoFLEX: Has 405 nm (Violet), 488 nm (Blue) and 633 nm (Red) lasers. Supports up to nine parameters and has a vSSC parameter for detection of nanoparticles at higher resolution. The CytoFlex configuration is customizable for specific experiments. The Cytoflex can perform absolute counts. Click here to access instrument configuration.
Automated Cell Counter
LunaII: The LunaII is an automated cell counter that supports brightfield cell counting. It has features to auto-focus, adjust light levels and capture a high-resolution image. You can create custom, cell-specific cell counting protocols or simply use default counting protocol suitable for most cell types.
|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|
|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|
Special filters: There are special filters available to detect fluorochromes that are not otherwise detectable by default configuration. Please contact us to see what special filters are available.
Primary Contact: David Rose, PhD
Manager, Flow Cytometry and Cell Sorting Facility
Office: (919) 513-6443
Flow Cytometry and Cell Sorting Facility
CVM Main Building, Room# B303
North Carolina State University – College of Veterinary Medicine
Raleigh, NC 27607
- Introductory Training
- Instrument Training
- Assisted Acquisition/Analysis
- Cell Sorting
- Analysis Software
- Experiment Design/Consultation
- Facility Access: Users who have been trained for independent operation of the analyzers will be given access to the online calendar system where they can schedule their appointments and will be given 24 hour access to the facility. Please contact core manager to schedule a training and request access to the facility.
- Instrument Training: First time users must go through a formal instrument training provided by the flow core. Instrument training by other fellow lab members is not acceptable, and such users will not be granted access to the instruments.
- First Time Users: For first time users, it is required to make an appointment with the core manager to discuss your proposed experiment to be sure the core facility has the capabilities to support your experimental needs.
- Cancellation: All appointments require 24 hour notice for cancellations. If you do not notify the core manager in writing, you will be charged.
- Billing policy: Billing will be based on either the scheduled time or the actual instrument usage time, whichever is greater.
- Scheduling Cell Sorting/Assisted Analysis: Users must schedule cell sorting and other facility-assisted appointments directly with the core manager, Javid Mohammed via email, as far in advance as possible.
- Biosafety: 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 core manager or EHS. Also, we are in the process of upgrading our facility to BSL2/2+. We will post the update here once the facility is upgraded to BSL2/2+.
- Listserv: 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.
- Acknowledgments: Please remember to acknowledge NCSU Flow Cytometry and Cell Sorting facility in all your publications resulting from the use of instrumentation and services at NCSU flow core. Where appropriate, please acknowledge individual members of the Flow Cytometry Core. Following text may be used to acknowledge facility use:
“Flow Cytometry experiments were performed in the Flow Cytometry and Cell Sorting facility at North Carolina State University- College of Veterinary Medicine.”
|Service||NCSU (per hour)||External Academic (per hour)||External Commercial (per hour)|
|LSRII unassisted||$ 56||$ 60||$ 70|
|CytoFLEX unassisted||$ 56||$ 60||$ 70|
|LSRII assisted (includes staff time)||$ 188||$ 190||$ 200|
|CytoFLEX assisted (includes staff time)||$ 188||$ 190||$ 200|
|MoFlo Setup (one time fee per sort)||$ 74||$ 80||$ 85|
|MoFlo Cell Sorting (includes staff time)||$ 215||$ 215||$ 220|
|Analysis Training||$ 160||$ 165||$ 175|
|Consultation||$ 70||$ 75||$ 80|
|Seahorse XF Mini- Customer operated||$ 61||$ 65||$ 75|
|Luna Cell Counter||Contact Us||Contact Us||Contact Us|
Flow Cytometry Tutorials:
- Fundamentals of Flow Cytometry (Video Tutorial)
- Introduction of FACS DIVA Software (Video Tutorial)
- Introduction to Flow Cytometry (Video Tutorial)
- Introduction to Fluorescence (Video Tutorial)
- Introduction to Flow Cytometry Data Analysis (Video Tutorial)
- Doublet Discrimination Tutorial – Video tutorial that covers different strategies to exclude doublets
- Cyto University: Free to ISAC members
Useful Flow Cytometry Links:
- The Flow Cytometry Network: A network to connect with Cytometry community
- BD Biosciences Flow Cytometry Tools
- Purdue University Cytometry Laboratories
- International Society for Advancement of Cytometry (ISAC)
- Biolegend Flow Cytometry Tools
- Optimized Multicolor Immunofluorescence Panels (OMIPs)
- Minimum Information about a Flow Cytometry Experiment
Flow Cytometry Tools:
- BD Spectrumviewer
- Thermo Fisher SpectraViewer
- Biolegend Spectra Analyzer
- FluoroFinder Panel Design Software
Flow Cytometry Protocols:
MoFlo-XDP Cell Sorter: Scheduling for cell sorting is only available though the flow cytometry core manager. Please contact us to discuss about your experimental needs and schedule a sorting.
LSRII and CytoFLEX: Scheduling for the LSRII and CytoFLEX is available through online calendar. Users are given access to the scheduler after completing required introductory and/or instrument training. Please contact the core manager to schedule introductory and analyzer training.