Published on 13 January 2015
Introduction to microfluidic stop flow
This guide describes how to generate a controlled flow inside a microfluidic chip and stop it completely (stop flow) by using an Elveflow® pressure control instrument (OB1) and a MUX Cross Chip.
Stopped-flow technology is frequently used to monitor rapid (bio)chemical reactions with high temporal resolution, e.g., in dynamic investigations of enzyme reactions, protein interactions, or molecular transport mechanisms.
First introduced in 1940 by Chance et al. and improved in 1964 by Gibson et al., stopped-flow technology is a highly versatile and convenient method for monitoring fast reactions.
The following video shows a stop flow experiment using fluorescent microspheres (sample) in poly-ethylene glycol (buffer medium) are used.
In order to perform a stop flow experiment, the sample must be miscible in the buffer medium and they must have the same density.
Three different setups are possible depending on the application and the number of inlets and outlets on the microfluidic chip.
The following setup allow a very good control of the sample in the microfluidic chip and is capable to stop the flow instantaneously but it is also possible to use two simpler setups with 3/2 valves using this application note.
Please note that in order to ensure a good zero flow performance, the sample solution has to be miscible and must have the same density as the buffer medium that fills the microfluidic system.
Applications
Results of a wide variety of applications can be improved by achieving stop flow in a microfluidic chip, from cell migration study due to chemotaxis (Yang et al., 2015) and precisely measure the performance of a microfluidic fuel cell (Cuevas-Muñiz et al. 2012) to long-term cell culture and detection (Sang et al. 2015).
This feature is also key in microscope-based projection photolithography and the formation of non-spherical particles (Dendukuri et al. 2007). Controlling perfectly how to stop the flow enables to decrease the number of imperfections on polymeric microparticles formed in situ and the automation increases the rate of formation. The advantage of the set-up including the MUX Cross Chip is mostly to work at very low volumes which is an interesting feature when one needs to reduce the mixing times, study reaction kinetics or minimize the volume in IR spectroscopy.
The MUX Cross Chip setup for stop flow
Principle
The MUX Cross Chip is a matrix of microfluidic valves that can be used for multiple applications, including stop flow but also phase injection sequences, flow focusing, etc.
Setting the pressure applied by the OB1 pressure controller at 0 mbar is not sufficient to totally stop instantaneously the flow as the pressure can take some time to stabilize, thus creating some residual flow at first.
A setup using a MUX Cross Chip has been developed to balance the pressure between the inlet and the outlet of the microfluidic chip, by applying the pressure from the OB1 flow controller. A syringe is used to perform a low volume injection experiment where the sample can be perfectly injected and controlled inside the chip. For example, it is possible to flow the sample both ways in the chip by applying successive positive and negative flows.
The sample is injected into the microfluidic chip using a syringe and a T-junction.
Be sure that all the cables and tubing are well connected to the Elveflow devices (USB cable, 24V DC, etc).
Perform leakage tests and remove any air bubble before starting the experiment to ensure a good flow regulation. This step is extremely important since air bubbles can contract or expand depending on the pressure applied, preventing the stop flow experiment to be instantaneous.

The MUX Cross Chip is a matrix of 4 x 4 valves. Connect any of the 4 inputs with any of the 4 outputs and open/close the valves using the software.
List of components

Fluidic 268 chip from microfluidic ChipShop

A syringe
Stop flow setup diagram

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Elveflow quick start guide
MAY THE FLOW BE WITH YOU
- Connect your OB1 pressure controller to an external pressure supply using pneumatic tubing, and to a computer using USB cable. For detailed instructions on OB1 pressure controller setup, please read the OB1 user guide.
- Plug the microfluidic reservoir to the OB1 pressure controller outlet. The Elveflow reservoirs connection instructions are covered by a specific guide (see Elveflow microfluidic reservoirs assembly instructions).
- For the feedback loop, connect a flow sensor to the OB1 with a wire. Then, connect it with tubes between the microfluidic reservoirs and the chip.
- Turn on the OB1 by pressing the power switch.
- Launch the Elveflow software. The Elveflow Smart Interface’s main features and options are covered in the Smart Interface guide. Please refer to those guides for a detailed description.
- Press Add instrument \ choose OB1 \ set as MK3+, set pressure channels if needed, give a name to the instrument and press OK to save changes. The OB1 flow controller should now be in the list of recognized devices.
- OB1 calibration is required for the first use. Please refer to the OB1 user guide.
- Add flow sensor: press Add sensor \ select flow sensor \ analog or digital \ max flow rate for the sensor, give a name for the sensor, select to which device and channel the sensor is connected and press OK to save the changes. The connected flow sensor should be in the list of recognized devices. For details refer to Microfluidic flow sensor user guide.
- Add MUX Cross Chip: press Add instrument \ select MUX Cross Chip and give a name to the instrument.
- Use the supplied 1/16” OD tubing to connect microfluidic reservoirs with the chip. Your MUX Cross Chip should be in the list of recognized devices.
- Open the OB1 and the MUX Cross Chip windows.
Stop flow experiment using a MUX Cross Chip
- Step 1 – Be sure that all the cables and tubing are well connected to the Elveflow devices (USB cable, 24V DC, etc). Fill all the tubes with the desired solution to remove any bubbles. (Put it in the reservoir. Open the Elveflow smart interface via the computer and select the OB1. Back on the software main window, set a pressure rate value in mbar (300 mbar for example) for the pertinent channel then click on the power sign button to start the flow). This step is extremely important since air bubbles can contract or expand depending on the pressure applied, preventing the stop flow experiment to be instantaneous.
- Step 2 – The majority of the bubbles that can be present inside the MUX Cross Chip should now be removed. To achieve this goal, the solution needs to be flowed from the input 1 to the output 1, then from the input 2 to the output 2, then from the input 3 to the output 3 and finally from the input 4 to the output 4. More steps can be added to remove the totality of the air inside the MUX Cross Chip if there are some difficulties to obtain a satisfactory stop flow. The MUX Cross Chip can be controlled via the ESI software to choose which valve to open. The following diagrams show how to perform this step.

- Step 3 – Close every valves of the MUX Cross Chip then follow the setup diagram above to assemble the experiment montage. The following valves can then be opened:

This configuration allows the injection of a little volume of sample into the T-junction when pushing the syringe plunger as the pressure at the inlet and outlet of the chip is equal to the pressure at the waste reservoir (atmosphere pressure). The syringe can then be removed and a cap can be put on the T-junction.
- Step 4 – A flow can now be created to push the sample into the chip by applying a pressure with the OB1 and by opening the following valves:

This configuration connects the inlet of the microfluidic chip with the OB1 and the outlet with the waste reservoir thus creating a positive flow. Applying a negative flow is also possible using the following configuration:

- Step 5 – Apply a stop flow by opening the following valves:

This configuration balances the pressure between the inlet and the outlet of the chip with good stability thanks to the pressure applied by the OB1, stopping the flow almost instantaneously.
Congratulations! You completely stopped the flow by using an Elveflow OB1 a MUX Cross Chip!
Hints & Tips
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Microfluidics knowledge
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