Viscosity describes the resistance of a liquid to flow and should be considered when designing your protocol in OneLab as it impacts the ability of the pipetting robot to properly transfer the liquid. This entails adjusting the pipetting parameters in OneLab by implementing commands specifically tailored for handling viscous solutions, thereby achieving improved pipetting performance and accurate transfer.

🍯 Viscous Solutions: glycerol, oil, bead slurry, non-ionic surfactants such as polysorbate or Tween 20.

Pipetting Recommendations (see full article here)

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

Parameter

Recommendations for Best Pipetting Viscous Solutions 🍯

Pipetting mode

Reverse pipetting;

Forward mode with air cushion control is an alternative

Pipetting speed

Slow to avoid the formation of air bubbles

Tip pre-wetting

Not recommended

Air cushion

Top air cushion if forward pipetting is used

Dispense tip position

Ideally at the liquid level or at the bottom especially if the consumable already contains other liquid

Pause before blow-out

Useful 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

Useful to remove residual liquid

Rinse after dispensing

Not possible

Repetitive dispensing

Not suitable

Tip type

Standard, ideally low retention

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 viscous 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

Air top cushion

Pipetting moving speed

Normal

Normal

Tip position at destination

w.r.t the liquid or the bottom

w.r.t the liquid or the bottom

Tip choice

Use tips w/ filter

No

No

Mixing at source ≡ Tip pre-wetting

No

No

Mixing at destination ≡ Rinse after dispensing

No

No

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

  • Perform a clean aspiration/dispensing

  • Increase the accuracy of the volume to be transferred

  • Reduce the chance of air bubble formation

Pipetting Performance Evaluation

As a viscous solution, we used Glycerol 75%. The images above demonstrate the pipetting output after dispensing glycerol 75% into a test tube containing distilled waters, using either the IDEAL (Reverse mode) or the ALTERNATIVE (Forward mode w/ air top cushion) 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. The droplet clinging at the tip end in (A) is normally removed by ensuring that the dispensing is properly performed with regard to the liquid surface or into the liquid.

To avoid any residual liquid from adhering to the pipette tip when following the liquid surface for dispensing, a liquid surface touch-off can be performed to ensure full, accurate liquid transfer. In OneLab, this entails slightly adjusting the tip position along the z-axis, enabling the release of any residual liquid adhering to the tip end by touching the surface of the liquid.

In (B), when using forward pipetting with an air top cushion, some residual liquid was present in the tip after dispensing. This residual volume can be minimized or even eliminated by ensuring that a pause is properly performed before the blow-out. A sufficient pause should allow the remaining liquid to flow down the interior of the tip and accumulate before the final purge.

Using one or the other of the two previous modes, 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.

In contrast, when using default forward pipetting settings with no custom control, the liquid transfer was inaccurate with sample loss due to incomplete, non-optimal dispensing.

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