Pipetting Viscous Solutions
Viscous Liquids: An Overview
To master pipetting, we need to understand that viscosity is the measure of a liquid’s resistance to flow. This can be easily seen when pipetting, by observing how quickly or slowly, the liquid flows when aspirating and dispensing. Pure water, and many aqueous buffers are low viscosity and flow easily. Honey, glycerol and polyethylene glycol are highly viscous and flow very slowly. Master mix is also viscous. There are additional properties such as vapor pressure, surface tension and contact angle that impact pipetting, but for this article, I’ll focus on the challenges unique to pipetting viscous liquids. So, how do we overcome these challenges?
Let’s explore a few tips and tricks.
Pipettes
Pipettes are commonplace in all laboratories, and for the most part air displacement pipettes are more typical in the lab setting. It should also be noted that manufacturers and calibration companies test and calibrate air displacement pipettes using low viscosity liquids such as water.
Positive displacement pipettes work better for viscous solutions because they have specialized tips that have a piston inside which is in direct contact with the sample. During dispensing, the piston wipes the inside the pipette tip and pushes all of the solution out of the pipette tip, regardless of viscosity.
The bottom line is that positive displacement pipettes work best for viscous solutions.
What if you don’t have a positive displacement pipette?
If a positive displacement pipette is not an option, don’t lose hope. Candie indicated that, “You can still achieve precision and accuracy using an air displacement pipette. When using an air displacement pipette, the reverse mode technique is the preferred method when pipetting viscous liquids.” With reverse mode pipetting you will aspirate the blow out volume but will not dispense it all out since you are only pushing the plunger to the first stop. Employing slow and steady techniques as listed above should also be executed. The following figure shows how to reverse pipette:
- If using an adjustable volume pipette, set it to the desired volume setting and load the tip on to the pipette.
- Hold the pipette vertically and depress the plunger completely. Go past the first stop and proceed to the second stop (often called the blowout stop).
- Immerse the tip in the liquid.
- Slowly release the plunger to full extension while the tip is immersed in the liquid.
- Remove the pipette from the liquid slowly. Dispense into the destination vessel by slowly pressing to the first stop (do not go past the first stop).
- Remove the pipette and tip slowly from the destination vessel. You should see a small volume of liquid remaining in the tip.
- Blow out this residual volume into a separate container and properly dispose of the used tip.
Pipette Tips
When it comes to pipette tips, it can be overwhelming to find the right tip for the job. In my discussion with Candie, we talked about concerns laboratories have with pipetting and how they overcome these challenges, especially when pipetting viscous liquids. She said, “I’ve heard that some labs cut the end of the pipette tip to increase the bore size of the tip, with the goal of improving pipetting but, when pipette tips are cut, the dimensions of the tip design changes and that results in inaccurate volumes being aspirated.” A better option is to use the specialized tips on the market designed specifically for challenging liquid types.
The take home message is that modifying the tip may appear to help with pipetting but will have adverse effects on accuracy. The best recommendation: Contact the pipette manufacturer to find the right pipette tip for pipetting viscous liquids.
Pipette Handling, Aspiration, and Dispense
When working with viscous liquids, pipette handling is just as important as using the right pipette and tips. I asked Candie about general handling practices when working with viscous liquid types. Candie added that, “When pipetting viscous liquids, pausing longer during aspiration, as well as dispensing, is needed.” It’s important to keep in mind that slow, steady movements when depressing and releasing the plunger will result in big accuracy and quality gains. It is also important to use steady motions while moving between aspiration and dispense so that there is no inadvertent damage to the pipette, loosening of the tip, getting extra solution stuck on the outside of the pipette tip, or shaking liquid out of the tip.
The key lesson here is that slow and steady movements will have a big effect on accuracy and precision.
Practice, practice, practice
As with mastering any skill, pipetting various liquid types takes practice. Each pipette and pipette tip will have a different feel. Practicing with various pipette types and tips will increase operator confidence, as well as improve pipetting precision and accuracy. You could also consider implementing an operator training program using viscous liquids. Glycerol, food oils, and even dish soap are great to use during training as they are challenging liquid types when it comes to pipetting.
Remember, practicing combined with feedback on accuracy and precision will improve viscous liquid handling confidence.
With the right tools, practice, and knowledge about your liquid type, you can achieve victory over viscous liquid pipetting challenges.
Additional Resources
About the Author
Breeann Bryan
Breeann Bryan is a dedicated laboratory professional with a LEAN Six Sigma Black Belt. Her background ranges from the bench to operational administration and project management. She is proud to share her knowledge and empower others to tackle their process improvement challenges, whether it’s troubleshooting data quality issues, finding out how to maximize efficiency in the lab, or keeping teams on task. She firmly believes that everyone deserves to have the right tools needed for the job.
