FACSAria The basic parts Overview Basic Parts of

FACSAria The basic parts

Overview

Basic Parts of the FACS Aria

Basic Parts of the FACS Aria Power switches Located at the left side Main Power turns on and off the Aria and the fluidics cart (via umbilical cable). The three additional switches turn on and off the power for the three different laser lines. Lasers can be turned off if not required for an experiment.

Lasers- On and Off The Manual recommends to keep the lasers off until the stream is on to protect the "dry" cuvette from direct laser light. However, if you do this, the instrument will disconnect from the software most of the times you turn the lasers on. . Solution: Start with all required lasers on! Keep the front cover open to protect the cuvette until the stream is turned on.

Basic Parts of the FACS Aria The circuit breaker needs only to be switched if there have been power surges in the laboratory. Switch first OFF, then back ON again. Do not use it as an additional power switch.

Basic Parts of the FACS Aria The fluidics cart provides air pressure and vacuum. It harbours the sheath and waste tanks and provides additional tanks for ethanol, water and cleaning solution.

Basic Parts of the FACS Aria

Basic Parts of the FACS Aria There should be no air bubbles in the filters. On new instruments or after filter changes, inspect the filters. If an air cushion is visible, vent the filter through the bleeder valve (power must be on, stream can be off!).

Basic Parts of the FACS Aria All tanks are pressure free. Thus, they can be opened and refilled „online“ during the instrument is running. However, disconnecting the fluidics lines from the ports requires to prime the lines after reconnection. Disconnect the lines only if you wish to clean the tanks. Do not disconnect the lines while the stream is on, because priming of the lines requires the stream to be off!

Basic Parts of the FACS Aria The fill level of the tanks is shown with small icons in the software. An error message will pop up to remind you to fill/empty the corresponding tank. Do not ignore this message! The sheath tank may run dry, hampering your sorts. The waste tank may overflow, requiring you to clean the lab. . .

Basic Parts of the FACS Aria The umbilical connection to the fluidics cart is properly configured during installation. The user can connect the temperature control and the AMS to the depicted ports.

Basic Parts of the FACS Aria The connection from the fluidics cart to the main instrument is configured during installation. Users may supply house air into the depicted port and switch off the internal compressor. The house air must at least supply 6 bar (90 psi)!

Basic Parts of the FACS Aria The sample port is software controlled. The sample chamber can be illuminated to check the fluidics level of the sample.

Basic Parts of the FACS Aria The system is delivered with different tube holders: • for 15 ml tubes • for 12 x 75 mm standard tubes • for 1 ml microtubes These holders are inserted into the sample port. Never insert tubes without the appropriate holder, as the sample station may become damaged.

Basic Parts of the FACS Aria The system is provided with three different collection tube holders: • for 15 ml tubes only two way sorting is possible (not enough deflection for four tubes) • for 12 x 75 mm standard tubes • for 1 ml microtubes

Basic Parts of the FACS Aria During operation of the sample port a red emergency stop button is lit. In case anything is hampering the operation of the chamber loading mechanics, pressing this button will: • Unload the tube • Stop the ACDU moving • Shut off the stream • Turn off the plate voltage • Protect the sample by closing the waste drawer • Put a warning message on the screen Thereafter the button can be reset by turning it to the left. Use with caution! The system has to be started up completely again!

Basic Parts of the FACS Aria Exchanging the sample lines will be explained during the trainings course.

Basic Parts of the FACS Aria Steering optics direct the laser light to the cuvette. The droplet breakoff point is observed by a dedicated camera.

Basic Parts of the FACS Aria Schematic drawing of the interrogation area. The cells are focussed in the flow cell, pass the interrogation point and thereafter enter the nozzle.

Basic Parts of the FACS Aria The three (depending on the configuration) laser intercepts are focussed onto three distinct collection fibers.

Basic Parts of the FACS Aria Each fiber leads to a dedicated detection block based on the Octagon principle. Each signal passes only one mirror. All mirrors are longpass and thus reflect light with lower wavelenghts. Due to this, an octagon can detect eight signals, seven colors plus the blue side scatter.

Basic Parts of the FACS Aria Signals from the red and violet lasers are directed to „Trigons“. At the moment there a few combinations for more than three colors fomr the red and violet lasers.

Basic Parts of the FACS Aria Schematics of the fluidics. After analysis the cells are either sorted or directed into the waste.

Basic Parts of the FACS Aria The sort chamber contains the plates and is closed during normal operation (Aerosol protection). The position of the red Accu. Drop laser can be adjusted by a micrometer dial.

Basic Parts of the FACS Aria Whenever the high voltage for the plates is on, a red warning light is lit. Take care when opening the sort chamber, as the plate voltage is about 6000 Volt!

Basic Parts of the FACS Aria The sort drawer protecs the samples and is only opened during sorts. The FACSAria detects clogs and closes the drawer automatically to protect your sorted samples. You can of course open and close it from the software manually.

Basic Parts of the FACS Aria The sample chamber can be set to a specified temperature to keep the sample cold or warm. It can only keep the sample at a given level, do not put a warm sample in and expect it to cool down to 4°C! Additionally, the chamber needs 30 minutes to reach the desired temperature!

Basic Parts of the FACS Aria There is a “bug” in the sample cooling: If you quit the software and start it the next morning, the sample cooling is still set to the desired temperature in the menu, however, the unit is NOT working. You have to turn it off and then select the desired temperature again in the software! You can feel the breeze outside of the exhaust fans.

Basic Parts of the FACS Aria The system has an inbuilt agitator to keep cells in suspension during sorting. However, bare in mind that this only keeps cells suspended, it is not a „Vortex“. Carefully resuspend all samples before introducing them to the sample chamber.