Here's some step-by-step advice on good pipetting technique:
- Make sure the pipette is set for the correct volume
- Ensure the tip is firmly attached
- Keep the pipette vertical when pipetting
- Slowly and smoothly depress the plunger to the correct point (the first stop position)
- Insert the tip in to the liquid to be pipetted. Ensure the tip is properly immersed in the liquid, but not touching/near any surfaces of the container
- Slowly and smoothly release the plunger back, keeping the pipette upright, and tip immersed, and making sure no bubbles form - you can try pre-wetting (see video linked below, around 3:00)
- Withdraw the pipette from the liquid
- Carefully dispense the liquid in to the desired container by steadily depressing the plunger to the second stop position (fully depressed), making sure all liquid is dispensed without spills or splashes.
"Put the tip in a 45-degree angle onto the wall of the container; then
start pushing down up until the first stop; then push through to the
second stop; keep the plunger depressed and remove pipette and check
whether the drop is in the container and pipette tip is empty" -
Christiaan
- Discard the tip using the release plunger
Also check you've got the right size/type of tip, and are using a suitable pipette (e.g. don't use a 10-100 µl pipette for 120 µl)
If this still causes incorrect measures then you could need to get your pipette calibrated.
Test a pipette by taking several samples. If the pipetted volume is consistent but incorrect (e.g. always 10% under the target) it suggests the pipette needs calibrating, it's probably not the technique that's wrong. If it is inconsistent (both under and over pipetting) then it's either technique or the pipette (bad/damaged pipettes can be inconsistent, poorly but good calibrated pipettes will more likely be consistent and only need calibrating).
Try using multiple pipettes, if they are each consistent but variance is among them (pipette 1 is always under by ~10%, pipette 2 is always over by 15%, pipette 3 is always over by ~2%...) then your technique is good, and you need better pipettes. Also get an experienced pipetter to check your technique and try the pipettes themselves.
Calibration can be done internally by lab techs or by external specialists, speak to your lab responsible person.
Also see this video which is well explained.