Foaming is the tendency of a liquid to build up foam in the presence of a surface-active material, such as surfactant solutions. It can be increased during pipetting by rough and excessive mixing or upon blow-out when air is introduced into the solution and therefore should be considered when designing your protocol in OneLab as it impacts the ability of the pipetting robot to properly transfer the liquid. Moreover, the resulting bubbles can have a negative impact on biochemical processes. This entails adjusting the pipetting parameters in OneLab by implementing commands specifically tailored for handling foaming solutions, thereby achieving improved pipetting performance and accurate transfer.

🧼 Foaming Liquids: protein-rich solutions (e.g. BSA-solution or cell culture medium), anionic surfactants such as sodium dodecyl sulfate or SDS.

Pipetting Recommendations (see full article here)

The table below summarizes the general recommendations for optimal manual pipetting of foaming solutions.

Parameter

Recommendations for Best Pipetting Foaming Liquids 🧼

Pipetting mode

Reverse pipetting;

Forward mode is an alternative

Pipetting speed

Slow to avoid foaming and bubble formation

Tip pre-wetting

Not recommended

Air cushion control

Not required

Dispense tip position

Ideally, at the liquid level or at the bottom especially if the consumable already contains other liquid; On the fly is feasible in some cases

Pause before a blowout

Helpful especially in forward pipetting mode when low-retention tips are not used

Pause after aspiration/dispensing

Recommended to ensure full deliver

Touch off after dispensing

Helpful to remove residual liquid

Rinse after dispensing

Rinse the tip after dispensing for a complete transfer when the forward pipetting mode is used

Repetitive dispensing

Suitable

Tip type

Standard, ideally low retention to avoid loss of sample or reagent

Tip filter

Not required

OneLab Pipetting Settings

The table below provides instructions on how to set up the OneLab software, following either the "IDEAL Setup" scheme or the "ALTERNATIVE Setup" approach, to ensure optimal pipetting results when handling foaming solutions while getting the best out of the pipetting system, whether you are using the guided Pipette+ system or the Andrew+ pipetting robot.

Parameter

IDEAL Setup

ALTERNATIVE Setup

Pipetting mode

Reverse

Forward

Blow-out

Disabled

Yes

Pause before blow-out

Disabled

Yes

Aspiration speed

Slow (1)

Slow

Dispensing speed

Slow

Slow

Air cushion

Disabled

No

Pipetting moving speed

Normal

Normal

Tip position at destination

w.r.t the liquid or the bottom; On-the-fly dispensing is feasible in some cases where volume accuracy is not critical (e.g. cell lysis using SDS-containing buffer)

w.r.t the liquid or the bottom; On-the-fly dispensing is feasible in some cases where volume accuracy is not critical (e.g. cell lysis using SDS-containing buffer)

Tip choice

Use tips w/ filter

No

No

Mixing at source ≡ Tip pre-wetting

No

No

Mixing at destination ≡ Rinse after dispensing

Yes (2), one to two times at a slow speed

Yes, one to two times at a slow speed

Mechanical arm motion speed

Normal

Normal

Custom tip position at the destination, customized dispensing height Liquid surface touch-off after dispensing

Apply a positive offset on the z-axis w.r.t the liquid surface to move the pipette tip slightly out of the liquid so as to ensure the clinging droplet or any residual liquid at the tip end is properly released

Apply a positive offset on the z-axis w.r.t the liquid surface to move the pipette tip slightly out of the liquid so as to ensure the clinging droplet or any residual liquid at the tip end is properly released

(1) Slow aspiration/dispensing speed = Low flow rate

Helps minimize the foaming effect and bubble formation.

(2) Rinsing the tip after dispensing for a complete transfer:

  1. After dispensing w.r.t to the liquid level, keep the tip in the solution

  2. Slowly perform one to two pipetting up/down cycles until the interior wall of the tip is clear

Pipetting Performance Evaluation

As a foaming solution, we used a cell lysis-like buffer containing detergent. The images above highlight the pipetting output after dispensing the foaming solution into a test tube containing distilled waters, using either the IDEAL (Reverse mode) or the ALTERNATIVE (Forward mode with tip rinsing) setup versus a default forward pipetting (without advanced control).

Based on the observations made during the test, the reverse pipetting delivered the most accurate transfer with no bubbles formed after dispensing.

On the other hand, The forward pipetting with tip rinsing after dispensing achieved a clean transfer with no residual liquid observed in the tip compared to the default forward pipetting without tip rinsing.

The bubble formation in both forward pipetting conditions may look similar, but the cause is different. In forward pipetting with tip rinsing, the bubbles were introduced by the one-time rinsing even at low speed (in OneLab, mixing up/down after dispensing in the destination) and the blow-out after the pause, whereas in the forward pipetting without tip rinsing, bubbles were generated by the normal dispensing speed and the abrupt blow-out without pause.

Using one or the other of the two pipetting solutions (ideal or alternative setup), one should always make sure that the pipette tip is correctly immersed in the liquid both at the source and destination, (generally, a few millimetres below the liquid's surface depending on the size of the tip) to allow proper pipetting of the desired volume and therefore improve accuracy. Immersing the tip too deeply in the liquid can increase the risk of liquid droplets clinging to the outside of the tip or liquid carryover, resulting in inaccurate pipetting. The tip immersion depth at the source and/or the destination vessel can be adjusted in OneLab through the "custom tip position" parameter.

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