Accurate pipetting is essential if you work on the bench. With inaccurate pipetting, you will have inaccurate stock solutions, your experiments will be unreproducible, and your assays will have such huge errors that comparing them would be worthless. But fortunately, there’s no need to get anxious—your reliable, precision micropipettes take care of that for you.
But handling these pipettes inappropriately leads to severe errors; therefore, the accuracy of your pipettes relies on you. You must maintain your pipettes, deeply understand their function, and practice good technique. Here’s our safe guide to accurate pipetting, which will bring your pipetting to the next level.
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Pre-Wet The Pipette Tip
Before aspirating a sample for delivery, aspirate and expel any sample liquid at least thrice; significant sample loss before delivery occurs due to evaporation within the tip. Pre-wetting increases humidity within the tip, decreasing sample evaporation variation. A lower volume in the first few samples can result from using the same tip to deliver multiple samples without pre-wetting. Working with volatile samples (i.e., organic solvents) increases the necessity to pre-wet.
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Work At Temperature Equilibrium
Before pipetting, allow liquids and equipment to be balanced to room temperature. The person or company that measured your pipette performed it at room temperature. If you’re performing at a different temperature (e.g., in a cold room), your pipette will not give out the displayed volumes.
Thermal expansion and shrinking can occur in the air space due to temperature differences. The influencing factor is liquid density after temperature equilibrium. Compared to room-temperature liquids, the cold liquid is more dense, and the hot liquid is less dense.
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Access The Pipette Tip For Droplets
Before dispensing, touch the reservoir’s side and wisely remove droplets outside the tip. When doing this, be cautious not to wick liquid out of the tip opening. After dispensing and before releasing the plunger, touch the tip to the container’s side and deliver any remaining liquid in the tip. Surface tension will help in drawing the remaining liquid from the tip.
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Based On The Liquid – Choose Forward Or Reverse PipettingÂ
Insert the plunger tip into the liquid, depress it to the first stop, and then release it to aspirate. To dispense the entire contents, remove the pipette from the liquid and press the plunger down to the second stop. Standard (or forward) mode pipetting provides greater accuracy and precision for all viscous or volatile liquids than reverse mode. Over-delivery frequently occurs in a reverse way. Therefore, it is advised to assess the impact of any potential over-delivery in the experiment and make changes as necessary.
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Pause Constantly After Aspiration
After aspirating the sample, leave a tip still in the liquid for about one second. It takes a minute for the liquid in the tip to complete moving after the plunger stops, so being unable to do so will cause the too-low volume. Making a pause constantly ensures correct aspiration.
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Remove The Pipette Straight From The Vessel.
Vertically hold the pipette and pull it straight out from the reservoir centre when aspirating liquid. This approach is especially vital when pipetting small volumes (less than 50µL – microlitre)—holding the pipette at an angle removed from the liquid changes the aspirated volume.
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Reduce Handling Of Pipette And Tip
By utilizing the finger rest, hold the pipette lightly. Ensure to return the pipette to the pipette stand between deliveries. Don’t use bare hands when handling pipette tips or reservoirs. Temperature equilibrium is interrupted by the body heat transferred during handling, which leads to variations in delivered volume.
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Use The Proper Immersion Depth.
Adequately immerse the tip below the meniscus before aspirating. Aspiration air occurs with too little immersion, particularly with large-volume pipettes. Too much immersion can cause liquid to cling to the tip outside. Limit aspiration by contacting the container bottom with the tip.
Conclusion
Here are the nine tips to improve your pipetting technique in 2022. We hope you are now aware of the pipetting approach and use them carefully by following these tips.
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