Director: Dr. Jon Oatley
Flow cytometry is a powerful tool for critically analyzing individual cells and other biological particles. Using this technology, expression of proteins on the surface of cells as well as simultaneous assessment of cell size and complexity can be investigated. A component of flow cytometry is fluorescent activated cell sorting (FACS) which allows for specific subsets of cells to be isolated in purity from a heterogeneous population. Moreover, isolation of single cells can be achieved using FACS allowing for critical examination of heterogeneity within diverse populations. The Center for Reproductive Biology at WSU has recently established a new FACS service core that is available to the general WSU community. The core contains two new state-of-the-art FACS machines manufactured by Sony Biotechnology that contain 4 lasers each. The SY3200 FACS machine is capable of 4 way color sorting and the SH800 is capable of 2-way color sorting. Both machines are available for use by the WSU community on a fee-for-service basis.
How to gain access to the FACS Core. The cell sorting instruments are housed in the basement of the new Veterinary and Biomedical Research Building (VBRB).
- Initiation of service is made by contacting the FACS core service manager Melissa Oatley (firstname.lastname@example.org). A pre-service meeting will be scheduled to discuss details of an individual investigators needs and technical considerations for setting up FACS experiments using the Core's specific machines.
- Sorting service: Conducted by the service manager based on the investigators pre-defined parameters.
- Service charges are established to cover costs of operating the machines and service contracts.
- SH800 Standard rate (CRB & Federal):First 100 hrs $58.00 (per PI, per year)
$116.00/hr after first 100Standard rate (CVM):$116.00/hrWSU-outside CVM & CRB:$145.00/hrOutside WSU:$156.60/hr
Minimum Charge: 30 minutes
Cancellation Policy: The minimum charge will be invoiced if an appointment is cancelled with less than 1 hour advance notice, 50% the minimum rate will be charged with less than 24 hours' notice.
- SY3200 Standard rate (CRB & Federal):First 100 hrs $59.50 (per PI, per year)
$119.00/hr after first 100Standard rate (CVM):$119.00/hrWSU-outside CVM & CRB:$148.75/hrOutside WSU:$160.65/hr
Minimum Charge: 1 hour
Cancellation Policy: The minimum charge will be invoiced if an appointment is cancelled with less than 3 hour advance notice, 50% the minimum rate will be charged with less than 24 hours' notice.
FACS Core Facts
Machines available for sorting:
- Sony SH800- 4 lasers (405, 488, 561, 638), 2-way sort capabilities
- Sony SY3200- 4 lasers (405, 488, 561, 642), 4-way sort capabilities
- If possible, appointments need to be made with a 1 week advance notice to ensure the availability of an instrument and sorting technician. Under special circumstances, an appointment can be made with at least a 48 hour advance notice. The clock for determining service fees starts at the agreed upon appointment time.
- Hours of operation are 8AM to 5PM Monday through Friday.
- Baring machine or operator error, the PI will be billed for ALL sorts regardless of whether the desired outcome is achieved. The core cannot guarantee a desired result if samples have not been properly processed for sorting or controls to set the sorting parameters are lacking.
Guidelines for Sample Preparation:
- Controls are vital to a successful sort. Negative controls (e.g cells incubated with normal isotype antibody or cells from wild-type litter mates when working with fluorescent reporters) are necessary for setting gates to collect the desired population.
- Samples must be a single cell suspension in a sorting media that has less than 1% FBS and little to no phenol red. Reagents are available from commercial sources for disassociation of tissues with a propensity for cell clumping. Note that cell viability should be assessed after generation of single cell suspension and prior to sorting.
- Sample tubes:
- SH800 – Samples can be processed in 0.5 ml, 1.5 ml, 5 ml, or 15 ml conical tubes
- SY3200 – Samples can only be processed in 5 ml round bottom tubes
- The ideal sample concentration will vary depending on the cell type. A good starting point is 4x106 cells per mL.
- Collection Tubes:
- Collection tubes are supplied by the investigator and should contain at least 250 uL of collection buffer. If post sort assessment is desired, the buffer should contain no more than 1% FBS.
- Tube sizes:
- SH800 – 5 mL round bottom and 15 mL conical tubes
- SY3200 – 5 mL round bottom and 15 or 50 mL conical tubes, and any size flat bottom well plate
The CRB will review all rates and accounts as per WSU Policies and Procedures.