Signup for a FACSAria run: please complete pre-experiment information (click here) and submit to kclass@umd.edu.

After approval notification, reserve time on the specified Oracle Calendar www.calendar.umd.edu, and:
1. select far right icon on the left side icon set, (the person/calendar icon)
2. then select resources, type in CLFS2102, click OK.  This will take you to the Flow Cytometry signup calendar.  You can view available slots and book appointments.

When booking appointments, please do so in 30-minute increments. 

• You will be charged for time signed up for unless canceled >24hrs in advance.

• Chargeback cost to UMD campus users will be:
  $25/hr sorting, $20/hr analysis only
  There will be a one-hour minimum charge due to time involved in setup and calibration of instrument.

• Samples will be run and data collected by the facility Director, Ken Class.  Frequent regular users who wish to run their own samples will be able to do so after undergoing one-on-one training and demonstrating proficiency in instrument operation.

Sample preparation:
All samples to be run on the FACSAria must be pre-filtered through no greater than 70um strainers prior to being run on the instrument.  (BD Falcon tubes, cat# 352235 are a preferred source. The flow lab has samples of these to try, but it will be necessary to eventually purchase your own).

• Samples to be delivered in 12x75 tubes.
• See page 3 regarding optimal cell concentrations.
• Required samples (controls/setup)
  Controls/compensation tubes.  All experiments must include:
• cells only control
• isotype controls
• Compensation controls, single stained tubes utilizing each dye in your experiment.  This is mandatory in multi-color (2 or more dyes) experiments since emission spectra from dyes overlap and non-specific fluorescence must be subtracted (compensated) from the various photodetectors.
• Sort samples are generally prepared in PBS+1%FBS.  Please contact Ken if alternative media is preferred/needed.
• Sort collection tubes (12x75) should contain approx 1ml of media + 20% serum + antibiotics (pen-strep, gentamycin), as this provides a cushion for the sorted cells and helps maintain sterility. Analysis-only samples not requiring viability can be fixed in 2% paraformaldehyde and run the following day, if necessary.

Examples of potential flow cytometric applications:
   Up to 9-color measurement of surface and/or internal antigens

• Cell cycle analysis (fixed or live cells)
• Measurement of fluorescent protein expression
• Apoptosis and cell death measurement
• Fluorescence Resonance Energy Transfer (FRET)
• Ion mobilization measurements (i.e. Calcium flux)
• Isolation of sub-populations of interest based on fluorescent characteristics

(Cell Sorting) into tubes or tissue culture plates.

Data analysis programs available: FACSDiva software, Flowjo (coming soon), Microsoft Office products.

FACSAria Operating Characteristics:

Excitation sources available and commonly used fluorochromes: 

Contact Ken at kclass@umd.edu regarding any additional dyes that are not mentioned in the table above.

*For sorting experiments, it is preferable to prep cells toward the higher end of the concentration range if possible.  Extra media can be brought to the sorting facility for diluting the sample if necessary. 

*For “analysis only” experiments, 0.5-5.0x106cells/ml are the optimal concentration regardless of nozzle tip size.