How to use the reverse pipetting technique to more accurately remove the protein solution

1. Press the button to the second stop.

2. Dip the tip into the liquid 1cm and slowly release the button to slide it back into place. This will fill the tip with liquid. Remove the tip from the liquid and contact the edge of the container to remove excess liquid.

3. Press the button smoothly to the first stop point when draining to the receiving container. Stay in this position. Some liquid will remain in the tip and cannot be discharged.

4. The liquid remaining in the tip can be blown back into the original solvent or discarded with the tip.

5. Release the button to the ready position.

Choosing the right pipette is also important for the precision of micropipetting. The ultra-blowing design of the Thermo Scientific F-series pipettes meets the need for micropipetting for precision. Thermo F series pipettes with less than 50μl of liquid range are designed with dual pistons, which increase air blowing capacity by 50%-60% compared to other common pipettes, so they are very clean when small volumes of liquid are blown out. Completely, greatly improving the accuracy of pipetting.

We used a Thermo Scientific Finnpipette F2 1-10 μl pipette with Thermo Scientific Finntip Flex 10 tips while pipetting 1% bovine serum albumin (BSA, Sigma A7030) using forward pipetting and reverse pipetting, respectively. Perform a pipetting accuracy test.

Figure 1 shows that when using the reverse pipetting technique, the change in pipetting is in a narrower range than using forward pipetting techniques.

Figure 2 shows the inaccuracy of using two pipetting methods. Inaccuracy is an estimate of the repeatability of pipetting. Reverse pipetting technology can reduce inaccuracy by 50% relative to forward pipetting techniques.

This is because the BSA solution contains a hydrophobic component that is easily adsorbed by the hydrophobic pipette tip wall. When using the forward pipetting technique, a small amount of liquid is likely to remain in the tip after each pipetting. This tendency increases the deviation between the volumes of the blown liquid, as the residual liquid accumulated in the tips when the pipetting is repeated may increase the amount of pipetting of the next pipetting. In the reverse pipetting technique, additional liquid is drawn into the tip. These additional liquids act like a reservoir which equalizes the amount of pipetting of the continuous pipetting. This reservoir also prevents air from entering the tip of the nozzle at the end of the blown liquid, which reduces the likelihood of liquid foaming. This makes reverse pipetting techniques especially useful when removing small liquid quantities. It can be seen that the selection of the Thermo Scientific F2 pipette and the reverse pipetting technique can improve the accuracy and repeatability of the pipetting protein solution.