10 Tips to improve pipetting technique
1. Pre-wet the pipette tip
Aspirate and expel sample liquid at least three times before aspirating a sample for delivery. Evaporation within the tip can cause a significant loss of sample before delivery. Prewetting increases the humidity within the tip, thus reducing variation in sample evaporation. Using the same tip (without prewetting) to deliver multiple samples results in lower volume for the first few samples. The need to pre-wet increases when working with volatile samples such as organic solvents.
2. Work at temperature equilibrium
Allow liquids and equipment to equilibrate to ambient temperature. The volume of sample delivered by air displacement pipettes varies with air pressure, relative humidity and vapour pressure of the liquid, all of which are temperature dependent. Working at a single, constant temperature minimizes this variation.3. Examine the tip before dispensing sample
Wipe the tip only if there’s liquid on the outside of the tip. Absorbent material rapidly carries sample from the tip if it contacts the tip opening. Unnecessary tip wiping increases the possibility of sample loss.
4. Use standard mode pipetting
Choose standard mode pipetting rather than “reverse mode” for all but viscous samples, if accurate and precise results are desired. In reverse mode pipetting, the plunger is depressed completely (past the first stop) to aspirate the sample and then depressed only to the first stop to deliver the sample. If reverse mode is used with normal aqueous fluids, the pipette tends to deliver more than the calibrated volume. On the other hand, using normal mode with viscous samples, especially when liquid is retained in the tip, results in under delivery.
- Good Pipetting Technique
5. Pause consistently after aspiration
Pause with the tip in the liquid for about one second after aspirating the sample. It takes a moment for the liquid in the tip to finish moving after the plunger stops, so failure to pause will cause the volume to be too low. Slow and even plunger release and a consistent, brief pause after aspiration minimizes this error.
6. Pull the pipette straight out
Pull the pipette straight out of the container after aspirating a sample. Do not touch the tip to the sides of the container. This technique is especially important when pipetting small volumes (<50µL). Surface tension effects cause the sample volumes to vary if the exit angles vary. Touching the tip against the container walls results in loss of sample.
7. Minimize handling of the pipette and tip
Set the pipette down between sample deliveries and avoid handling the tip. Body heat transferred to equipment during handling disrupts temperature equilibrium. As explained in Tip no. 2, the volume of sample delivered varies with temperature.
8. Immerse the tip to the proper depth
During sample aspiration, immerse the tip 2–5mm below the meniscus and well clear of the container walls and bottom. Inserting the tip too deep into the liquid causes excess droplets to cling to the outside of the tip. Pressing or resting the tip against the container walls or bottom restricts entry of the sample.
9. Use the correct pipette tip
For accurate volume delivery, choose a tip that’s designed for use with the type of pipette being employed and securely attach it. Mismatching a tip and pipette, or using poor quality tips can result in an inadequate seal between the pipette and tip. Quality tips are flexible and have thin walls, providing airtight seals and dependable sample delivery.
10. Use consistent plunger pressure and speed
Depress and release the plunger smoothly and with consistent pressure and speed when dispensing each sample. Pipettes, like all precision instruments, produce more reproducible results when operated with attention to detail and proper technique.
For more informaton on the full range of Camlab’s pipettes and tips please visit www.camlab.co.uk
In: 1. News · Tagged with: pi petting technique, pipettes