Aria, sequential perfusion system

[OAR]

    Automated system for sequential perfusion experiments

    Fluigent introduces Aria, our automated solution for cellular perfusion or timed injection protocols. Aria allows one to automate delivery up to 10 different solutions into a chamber or microfluidic chip by following user defined protocols.

    Main benefits
    • Automation

      Long-term perfusion studies

    • Biocompatible
      Biocompatible

      Contamination-free

    • Save time

      Automated protocols

    • Easy to use

      All in one instrument and software

    • Adaptable

      Fits any experimental design

    Features

    Deliver up to 10 solutions

    Aria covers any applications as the volumes injected range from 40µL to hundreds of mL delivered over several days. To minimize carryover between successive solutions, the tubing can be cleared. The user is informed of the delivery time of each solution thanks to Aria’s intelligent software. The software also notifies the user of the minimum volume required in each reservoir to run their protocol

    Automate any protocol

    Aria comes with intuitive software that enables to design protocols in a few clicks. Incubation time, flow rate, volume dispensed, are all parameters that can be easily set by the operator for each step of his protocol. Protocols are recorded and can be shared among users. Flow rate and pressure are recorded for each experiment

    Reduce variability

    Aria drastically reduces variability down to 0,5% between experiments compared to 5,1% intra-operator variability and 8,1% inter-operator variability using a pipette

    Preserve sample integrity

    Contamination free: the sample is delivered at the beginning of the experiment and is not touched during the protocol minimizing contamination risks from manual operations. Controlled, smooth flow preserves samples compared to traditional pipetting.

    Ideal for imaging studies

    Aria can synchronize with various microscopes through the use of TTL signaling. Aria can send and receive TTL signals for example to start an imaging cycle, or to resume the Aria protocol when an imaging cycle is completed. Aria’s software comes with a dark mode to work in an imaging room. Aria is equipped with LED’S to facilitate use in dark areas

    Why Aria?

    Applications like DNA hybridization, high resolution spatial transcriptomics, DNA paint, or tissue profiling require high resolution microscopy as they target nanometric structures inside the cells.

    For each of these applications, the sample is successively incubated with specific probes to bind to a target inside the cells. Between various probes, a sequence of washing steps is performed to remove the previous probe and avoid contamination with the consecutive one.

    The power of this technique compared to traditional immunolabeling is that the number of targets per sample is not limited to the four traditional colours of the microscope as DNA can be easily detached from their target as opposed to antibody that remain strongly attached when bounded.

    Reduce experimental error

    Manually changing solutions from washing buffers to probes on the microscope comes with some challenges:

    • A touch of a pipette tip can displace the dish and recorded positions
    • Samples can be flushed away during pipetting
    • The sample can get contaminated
    • Liquid can spill over the microscope
    • Errors in the sequence of solutions due to the number of steps

    In addition, successive solution replacement can be long (up to 4-5 days for an experiment), tedious and requires the physical presence of the scientist.

    Automate protocols

    Aria is the only instrument that can automate the delivery of up to 10 different solutions to a flow cell. It is the perfect compromise between manual pipetting and all-in-one system that are dedicated to one specific application, integrate a microscope, specific chip type and a given set of solutions: Any protocol with multiple solution delivery can be automated, saving the scientist both time and reducing variability between experiments compared to manual procedures.

    • Aria is straightforward and ready to use. No specific training is required, and no time is lost trouble shooting incidents or manually priming solutions before the experiment as is the case with home-made systems.
    • Compared to fully integrated systems that are bound to one chip type and a set of solutions for a given application, Aria offers flexibility. Aria can automate different applications in one lab as it is compatible with any chip and any type of reagent. Aria is transportable and can be easily moved between microscopes.
    • Aria is a cost-effective alternative to an all-in-one system. It is perfect to investigate and set all the parameters to fully optimize imaging protocols.


    “I got to test Aria injection system in my research project in a collaboration with Fluigent. More precisely, Aria injection system helped me automatize the capture process and immunostaining of breast cancer cells under a very precise and controlled flow rate. The software interface is so user-friendly that I was being able to follow in real-time the progression of my experiment. The amazing part is that ARIA even calculated the total amount of time required for each step and helped me avoid the waiting time in front of my setup! It made my experiment go as smoothly as possible.”

    Emile Lakis / Curie Institute, IPGG / Paris

    Choose the version that best meets your needs

    Single output version

    Perfect for high quality multiplexed imaging experiments.

    Aria is the perfect compromise between manual pipetting and all-in-one system that are dedicated to one specific application, integrate a microscope, specific chip type and a given set of solutions: Any protocol with multiple solution delivery can be automated, saving the scientist both time and reducing variability between experiments compared to manual procedures.

    Serial output version (perfuse up to 9 samples)

    Perfect benchtop workstation to automate staining or any routine protocols with multiple fluid delivery.

    Aria is designed to fully automate your protocols, gain productivity, reliability, reduce manual operation and save time. Aria is the only workstation on the market compatible with any perfusion chamber.

    Specifications

    PERFORMANCE

    Flow rate controlOver the range of 40 µL/min to 1 mL/min for water
    Pressure control of flow rateto a maximum of 2 bar
    Valvesten position switching valve – two position switching valve (2)
    Fluid reservoirs (8)15 mL standard, 2 mL available
    Flushing solution reservoir (2)100 mL
    TubingFEP with OD of 1/16 inch and ID of 250 µm
    Wetted surfacesPolypropylene, FEP, Glass, PEEK
    Compressed air sourceRequires non-corrosive compressed air (lab line, gas
    cylinder, compressor or Fluigent FLPG)
    PC specificationsWindows 7 or higher

    HARDWARE SPECIFICATIONS

    Dimensions382 mm x 240 mm x 265 mm
    Weight9 kg

    ELECTRICAL SPECIFICATIONS

    Power supply voltage24V DC

    CHEMICAL COMPATIBILITY

    Gas compatibilityDry, oil-free gas, air, any non corrosive or non explosive gas
    Liquid compatibilityAqueous solutions only

    Aria software

    Control in real-time, protocol automation, data record and export

    Episode 1

    How to plug an Aria


    Episode 2

    Connect to set the fluidic path


    Episode 3

    How to fill up the reservoir


    Episode 4

    How to use Aria manually


    Episode 5

    Perform a calibration with Aria


    Episode 6

    Global handling of Aria


    Episode 7

    Software presentation

    To save time and minimize reagents consumption, Aria loads its internal and external tubing with the successive solutions to inject in the chip.

    Working principle:

    If the protocol commands to inject 40 µL of solution A, incubate for 1 hour, inject 50 µL of solution B, incubate for 1 hour then wash with buffer for 5 minutes and the total volume of tubing ( internal + external) is of 300 µL Aria will prefill tubing as follows:

    As a consequence, calibration of the tubing length and related volume is necessary.

    Here is a description of all the functions available to optimize protocol writing in Aria

    Calibration

    Calibration is automated and the software assists the user to determine the total volume of his step up (internal volume + L1+L2). Calibration values are recorded for future experiments.

    Prefill

    Initially all tubing inside Aria are dry. The “Prefill” function is recommended as a first step to load all solutions inside Aria without injecting air in the chip or chamber. This function can be deactivated if the user performed the loading manually. Fluigent strongly recommends using this function.

    Perfusion/injection

    Perfusion can be set in terms of volume (ex: step 1) or duration (ex: step 2) of injection.

    The user selects the reservoir of the solution to inject (ex: reservoir 1), sets the flow rate (ex: 100 µL/min) at which the solution will enter the flow cell and the volume (ex: 100 µL) or duration (ex: 1min) of injection. The flow rate  range is from 40 µL/min to 1 mL/min. The software automatically calculates the time at which the solution should enter the flow cell.

    Incubation/wait

    Incubation time can be easily set by entering the duration of the incubation (ex: 1h30) in the ‘wait’ function.

    The ‘wait for user’ function is a variation of the ‘wait’ function. It is particularly useful if the user has to perform a manual operation like preparing cells before injection. The system waits until the user notifies it can proceed to the next step. In absence of notification by the user, the system proceeds to the next steps after 12 h.

    Clear tubing

    To prevent contamination between successive injections, the tubing can be cleared with buffer using ‘Flush tubing’ function. The user selects the reservoir containing the wash solution (ex: reservoir 10) and the flow rate at which the tubing is flushed (ex: 100 µL/min).

    Aria pushes all the residual fluids contained in the L1 tubing to waste, and fills the L1 with wash solution. Buffer contained in L1 will also be directed to the waste. This operation does not involve flow to the  chip as neither residual fluids nor buffer enter the chip. However, the residual fluid contained in L2 will not be cleared. For this reason, Fluigent  recommend the user to keep L2 as short as possible.

    External synchronization

    Aria is equipped with TTL signalling and can send and receive external TTL signals.

    • Sending TTL

    Each step is flanked by two bells () : at the front and back of the settings (see below for ‘volume injection step).

    By clicking on each of both of the bells they get activated (). Aria will then send a ttl signal either when the step starts (example below) or ends or both.

    Fluigent recommends using this function to synchronize perfusion and imaging.

    • Receiving TTL

    The function ‘wait for TTL’ puts Aria on hold until it gets an external TTL signalling to proceed with the next steps of the protocol. If the Aria does not get any signalling it automatically proceeds with the next steps after 12h.

    Minimum volume reservoir

    Aria is designed to minimize reagent consumption. A small additional volume of liquid (36µL) is necessary to preload the system and to ensure that some residual liquid is still present in reservoir after injection to prevent delivery of air to the system.

    As the user writes his protocol, the software calculates the minimum volume that should be placed in each reservoir to successfully run the protocol.

    Write, load, save sequence

    To design a new sequence, click on .

    Before running a sequence, the software will automatically ask the user to save it. All saved sequences are accessible and can be loaded by clicking on .

    Sequences can be saved at any time by the user by clicking on the saved data .

    Recorded data

    For each experiment, Aria automatically records and saves the flow rate, pressure, the reservoir from which the solution is withdrawn if solution is delivered to the chip or waste. All data is accessible by clicking on .

    Publications

    Radtke, A.  <https://arxiv.org/abs/2107.11364> et al. (2021)IBEX: An open and extensible method for high content multiplex imaging of diverse tissues, In Press

    Huang, K.  <https://bio-protocol.org/e4128?p=51&way=207> et al. (2021). A Novel Method to Map Small RNAs with High Resolution. Bio-protocol 11(16): e4128. DOI: 10.21769/BioProtoc.4128.

    Presentation

    Radtke, A. <https://drive.google.com/file/d/1QkJSvtH2oAUFrssg-Sp7jp5XQQysuDP9/view> et al. (2021)IBEX: An open and extensible method for high content multiplex imaging of diverse tissues, Presented at CZI HCA network and HuBMAP consortium