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Reverse pipetting is a technique to dispense a measured quantity of liquid by means of
In reverse pipetting, the volume aspirated to the tip is bigger than the volume delivered to the receiving vessel.
The excess of volume compensates for the liquid that remains as film inside the tip during dispensing. Liquid aspiration is achieved by creating vacuum by means of the vertical travel of a metal or ceramic piston within an airtight sleeve. As the piston moves upward, driven by the depression of the plunger, a vacuum is created in the space left vacant by the piston. Air from the tip rises to fill the space left vacant, and the tip air is then replaced by the liquid, which is drawn up into the tip and thus available for transport and dispensing elsewhere.
When the pipette knob is pressed on an
These pipettes are capable of being very precise and accurate. However, since they rely on air displacement, they are subject to inaccuracies caused by the changing environment, particularly temperature and user technique. For these reasons this equipment must be carefully maintained and calibrated, and users must be trained to exercise correct and consistent technique.
Alternative solutions to improve reproducibility and accuracy of manual pipetting operations are based on anthropomorphic liquid handling robots capable to handle air displacement pipettes.
Guide to pipettingGood laboratory pipetting guideThermo Scientific Pipetting GuideWikilecturesPipetting Viscous SolutionsTips for Proper PipettingHow to pipette correctly - 10 tips
Andrew Alliance websiteGilson websiteINTEGRA Biosciences websiteThermo ScientificSocorexBiohit
air displacement pipette
Piston-driven air displacement pipettes are a type of micropipette, which are tools to handle volumes of liquid in the microliter scale. They are more commonly used in biology and biochemistry, and less commonly in chemistry; the equipment is su ...
. The technique is mainly recommended for solutions with a high viscosity
Viscosity is a measure of a fluid's rate-dependent drag (physics), resistance to a change in shape or to movement of its neighboring portions relative to one another. For liquids, it corresponds to the informal concept of ''thickness''; for e ...
or a tendency to foam: as it reduces the risk of splashing, foam or bubble formation. Reverse pipetting is more precise in dispensing small volumes of liquids containing proteins and biological solutions compared to forward pipetting Forward pipetting is a technique to dispense a measured quantity of liquid by means of air displacement pipette. The technique is mainly recommended for aqueous solutions, such as buffers, or diluted acids or alkalis. In case of solutions with a hi ...
, which is mostly used for aqueous solutions, such as buffers, diluted acids or alkalis.
How it works
In reverse pipetting, the volume aspirated to the tip is bigger than the volume delivered to the receiving vessel.
The excess of volume compensates for the liquid that remains as film inside the tip during dispensing. Liquid aspiration is achieved by creating vacuum by means of the vertical travel of a metal or ceramic piston within an airtight sleeve. As the piston moves upward, driven by the depression of the plunger, a vacuum is created in the space left vacant by the piston. Air from the tip rises to fill the space left vacant, and the tip air is then replaced by the liquid, which is drawn up into the tip and thus available for transport and dispensing elsewhere.
When the pipette knob is pressed on an air displacement pipette
Piston-driven air displacement pipettes are a type of micropipette, which are tools to handle volumes of liquid in the microliter scale. They are more commonly used in biology and biochemistry, and less commonly in chemistry; the equipment is su ...
, the piston inside the instrument moves down to let air out. Air is displaced by the piston. The volume of air displaced is equivalent to the volume of liquid aspirated.These pipettes are capable of being very precise and accurate. However, since they rely on air displacement, they are subject to inaccuracies caused by the changing environment, particularly temperature and user technique. For these reasons this equipment must be carefully maintained and calibrated, and users must be trained to exercise correct and consistent technique.
Alternative solutions to improve reproducibility and accuracy of manual pipetting operations are based on anthropomorphic liquid handling robots capable to handle air displacement pipettes.
Technique
Reverse pipetting with manual pipettes
Reverse pipetting with electronic pipettes
Some electronic pipettes have a mode for reverse pipetting which allows to set the following parameters: # Desired volume # Excess volume (about 3-5 % of the nominal volume of the pipette) # Specify whether the excess volume should be discarded or included in the next aspiration # Aspiration speed # Dispense speedReferences
Sources
Guide to pipetting
External links
Andrew Alliance website
See also
* Automated pipetting system * Liquid Handling Robot *Pipette
A pipette (sometimes spelled as pipet) is a type of laboratory tool commonly used in chemistry and biology to transport a measured volume of liquid, often as a media dispenser. Pipettes come in several designs for various purposes with differing ...
*Air displacement pipette
Piston-driven air displacement pipettes are a type of micropipette, which are tools to handle volumes of liquid in the microliter scale. They are more commonly used in biology and biochemistry, and less commonly in chemistry; the equipment is su ...
*Forward pipetting Forward pipetting is a technique to dispense a measured quantity of liquid by means of air displacement pipette. The technique is mainly recommended for aqueous solutions, such as buffers, or diluted acids or alkalis. In case of solutions with a hi ...
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Laboratory equipment