How do I pipette volatile liquids?

Introduction

A volatile liquid is any liquid that evaporates quickly. They often have low surface tension and may drip from pipette tips… a lot. Ethanol is a commonly used example of a volatile liquid. Researchers will often struggle to pipette volatile liquids until they learn the know-how.

1. Do it quickly

The faster you complete the liquid transfer and get those caps back on, there’s less time for liquid loss from evaporation or drippage.

2. Take up all the space

Even when they are inside the pipette tip, volatile liquids will evaporate into any space above the top liquid line. This results in a small amount of liquid loss to gas and increased vapour pressure within the tip. Vapour pressure is exerted back onto the liquid causing even more dripping. This is exacerbated by higher temperatures. To combat this you should use the right size pipette tip for your volume, ensuring that the majority of the tip contains liquid. 

3. Lids on – ASAP

It seems obvious, but it is the most important part. Keep lids/caps on your tubes for as long as possible to prevent premature evaporation. This won’t always be possible for an automated liquid handler, but some have options for a manual step/intervention in the workflow. You can use this to remove and replace lids and caps mid-workflow. 

4. Pipette at a lower temperature

For a manual pipette, this will simply mean finding your nearest cold room (if you have one) and doing it there. Using a cold block is also a valid option. For an automated liquid handler, often specialist cold blocks or cooling equipment is required that are compatible with your setup. However, if you’re only going to do this once, it’s more expensive. An alternative option would be to just pick up the entire unit and take it to the walk-in fridge as you did with the manual pipette (just double check your standard operating conditions). Lowering the temperature will prevent liquid loss through evaporation and also reduce the fluidity, ultimately increasing its adherence to the pipette tip. This may help battle against dripping as well; every little helps. 

5. Use an air gap

When using an automated liquid handler, it’s possible to add an air gap beneath the aspirated liquid. This creates a barrier/buffer zone between the volatile liquid and the bore of the tip. The liquid may still struggle to adhere to the inside of the pipette tip but there is greater distance between the bottom liquid level and the bore before the liquid drips.

6. Additional liquid

To compensate for the inevitable dripping, use reverse pipetting. This means to aspirate more volume than will be dispensed. For a manual pipette: press down to the second position and aspirate to a relaxed position then dispense to the first position. For an automated pipette: set the aspirate volume greater than the dispense. This can also benefit point 2, as you can fill the entire pipette tip with liquid and still dispense a small amount unlike the manual pipette. Any excess can be discarded or returned to source.

7. Reduce the concentration

Water has the highest surface tension of the non-metallic liquids. By reducing the concentration of the volatile liquid before pipetting, you are increasing the surface tension (liquid will bind itself together) which will reduce the amount of dripping.

8. Increase the volume

This is purely a numbers game. For example 1 μL liquid loss in a 50 μL transfer won’t make a huge difference in the long run, but 1 μL loss in a 10 μL transfer is a significantly greater degree of error.

Summary

As always, the best way to ensure accurate and reliable liquid transfer is to use regularly calibrated and cleaned pipettes with pipette tips from a reputable supplier. Good equipment is the key. Singer Instruments has 90 years of experience in high quality lab equipment and excellent user support.