Fees effective: June 1, 2015

These rates are for Arizona Universities. For external pricing, please contact the Facility Manager or the ARL Business Office (520-621-4064).

Acquisition has priority over data reduction.

Cell-sorting users will be charged $123.00 for setup, and then $123.00 per hour for sorting prorated to 15 minute increments.

Consultation services and the use of support equipment are available free of charge.

External customers: Please contact the Core Facility Manager.

Individual consultation services are available to discuss applications and methodologies specific to your project.

Contact the Facility Manager to set up an appointment.

There is also a small library of books and reference materials which are available for use.

Subpopulations of cells or particles that have been resolved by fluorescence or physical parameters can be isolated for further experimentation by using the instrument's cell sorting capability.

A piezoelectric quartz crystal connected to the nozzle causes the cell stream to oscillate and form droplets. The droplets are formed at a high enough frequency so that any given droplet will contain only one cell. When a cell meets the criteria defined by sort "windows", the sorting electronics system places a charge on the cell stream at exactly the time when the droplet containing the cell of interest is breaking off from the stream. The droplet passes between two metal plates that have a very high voltage.

The resulting magnetic field forces the charged droplet to be deflected toward the plate with the opposite charge. By placing a collection vessel in the deflected droplet's path, cell populations up to 99% purity can be collected at rates of approximately one thousand per second. The current flow cytometer can sort four populations at the same time.

The BD FACSAria cell sorter is operated by Cytometry Core Facility personnel only. Please contact the Core Facility Manager to discuss your cell sorting needs.

This form must be filled out and submitted to the Facility Manager prior to starting any new sorting projects involving live human or pathogenic samples.

Flow cytometry is a powerful and effective tool for analyzing the structural and functional characteristics of cells or particles in suspension. A test tube containing a single cell suspension is placed on the instrument where the sample is pushed by air under pressure through tubing where is is injected into a nozzle. The nozzle, containing sheath fluid under pressure, hydronamically focuses the cells so that they exit the nozzle in single file down the center of the stream resulting in laminar flow. The flow rate can be controlled by adjusting the differential pressure between the sheath fluid and the sample injection pressure. Laser light is then focused onto the stream where each cell is excited as it passes through the laser beam.

Photo multiplier tubes (PMT's) are detectors which collect the photon emissions from each "event" and convert them to analog voltages. The analog signals are then digitized by analog to digital converters (ADC's) and recorded as data files. Optical filters are placed before the detectors so that only wavelengths of light corresponding to specific fluorochrome emissions are collected by each detector (e.g. FITC emits in the green region therefore a 30nm bypass filter centered at 520nm could be used to collect light from this fluorochrome). Light scattered at the same wavelength and direction as the laser light, primarily from the surface of the cell, correlates with relative cell size (Forward Angle Light Scatter) while light scattered 90 degrees to the laser (Side Scatter) usually from internal structures, correlates with granularity. By correlating these two parameters, one can discriminate subpopulations of cells in peripheral blood samples, for example. Signals corresponding to cell debris or cell aggregates can also be detected and excluded from analysis on the basis of forward and side scatter.

Staining cells with multiple fluorochromes conjugated to antibodies or fluorochromes directed at other specific targets such as DNA, cytokines, or other proteins distinguishes cell subpopulations which can be quantified. Data is displayed and analyzed using histograms or two-dimensional dot plots on a computer system.

The Cytometry Core Facility has two Analyzers available for use: a two-laser BD FACSCanto II and a five-laser BD LSR II

All services being with a consultation with the Facility Staff.

Appointments for the FACSAria and the LSR II must be made by directly contacting the Facility Manager.

For access to the FACSCanto II after your initial consultation, complete and submit the New User Registration form.

1. Use your email login (the first part of your email address before the "@") as the Login Name.
2. Click Submit when you have provided all the information required. Access will not be approved until all information is complete.
3. Contact the Facility Manager when you've submitted the access request and we will respond by email with your approval.
4. Once access has been approved, you can schedule appointments for the FACSCanto II using the scheduling calendar.
5. Samples for the FACSCanto II optimally should contain 1x10⁶ cells/ml (300,000 cells minimum). For tubes that meet these requirements, allow on average 3 minutes per tube (5 minutes for cell cycle analysis) for most samples.

Experiments that look for rare events will require more time per sample.

Remember, the fewer the cells, the longer the time required for data acquisition per tube.