A micropipette is a laboratory instrument designed for the precise measurement and transfer of minute volumes of liquid, typically in the microliter (\(\mu L\)) range. These volumes, often less than a thousandth of a milliliter, require specialized tools for accurate handling. The reliability of results in modern scientific disciplines, including molecular biology, clinical diagnostics, and drug discovery, depends heavily on this precision. Minor errors in liquid handling can lead to significant variations in experimental outcomes, making the correct use of the micropipette fundamental to scientific validity.
The Anatomy of a Micropipette
The core of a standard air-displacement micropipette is a piston-driven system that uses an air cushion to aspirate and dispense fluid. This design prevents the internal mechanical components from contacting the liquid sample, maintaining the instrument’s integrity and preventing cross-contamination. The main operational control is the plunger button, which manages the aspiration and dispensing steps.
The plunger features two distinct resistance points: the first stop and the second stop. The first stop corresponds to the set volume and is used to expel air before aspiration and to dispense the measured liquid. Pushing the plunger to the second stop engages an extra volume of travel, used as a “blowout” to ensure residual liquid is fully expelled from the disposable tip.
Volume selection is managed by a volume adjustment dial, often integrated into the plunger mechanism, which sets the desired volume displayed in the indicator window. The lower end of the pipette terminates in the tip cone, onto which a disposable plastic tip is securely seated. After use, the tip ejector button safely removes the disposable tip, preventing the operator from touching potentially contaminated plastic.
Different Pipetting Mechanisms and Designs
Micropipettes utilize different internal mechanisms for accurate liquid transfer, primarily categorized as air displacement or positive displacement. Air displacement pipettes are the most common type, often considered the laboratory workhorse for handling aqueous solutions. They rely on a piston displacing a volume of air, which then draws liquid into the tip.
The air cushion separating the piston from the liquid makes air displacement pipettes susceptible to inaccuracies when working with “difficult” liquids. Highly volatile solvents can evaporate into the air cushion, leading to under-delivery of the measured volume. Viscous liquids, such as glycerol or oils, also require a different mechanism because they adhere strongly to the inner tip wall, resulting in inaccurate dispensed volumes.
Positive displacement pipettes overcome these issues by integrating the piston directly into the disposable tip. This design eliminates the air cushion entirely, ensuring the piston is in direct contact with the liquid sample. This allows for reproducible results when handling challenging substances such as volatile, dense, or high-viscosity fluids.
Micropipettes are also categorized by structural variations, such as fixed or adjustable volume models. Fixed volume pipettes dispense a single, specific volume repeatedly, benefiting routine, high-volume workflows requiring consistency. Adjustable volume pipettes offer flexibility, allowing the user to select any volume within the instrument’s designated range, ideal for research laboratories.
Pipettes are available as single-channel models for individual liquid transfers or as multichannel versions, which typically contain 8 or 12 dispensing heads. Multichannel pipettes significantly increase efficiency for high-throughput applications, such as filling a 96-well microplate for diagnostic tests like ELISA or PCR.
Essential Techniques for Accurate Pipetting
Achieving accuracy with a micropipette depends heavily on the operator’s technique, beginning with correct instrument preparation. Before drawing up a liquid, the disposable tip should be pre-wetted two or three times by aspirating and dispensing the sample liquid. This step equilibrates the air temperature and humidity inside the tip, reducing evaporation and minimizing liquid clinging to the tip surface.
The standard, or forward, pipetting technique is used for most aqueous solutions. It begins by depressing the plunger smoothly to the first stop. The tip is immersed just a few millimeters beneath the liquid surface, and the plunger is then released slowly and evenly to aspirate the measured volume. Releasing the plunger too quickly can cause air bubbles or splash liquid into the pipette cone, risking internal contamination.
To dispense the liquid, the tip is generally placed against the inner wall of the receiving vessel at a slight angle. The plunger is depressed slowly to the first stop to dispense the set volume, and after a brief pause, it is pushed all the way to the second stop. This final movement is the blowout, which clears any remaining liquid film from the tip, ensuring the full volume is delivered. The tip must be removed from the liquid while the plunger is still held at the second stop to prevent re-aspiration of the dispensed fluid.
Reverse pipetting is favored when handling viscous or volatile liquids. This technique starts by depressing the plunger all the way to the second stop to aspirate a volume larger than the set amount. When dispensing, the plunger is pushed only to the first stop, leaving the excess liquid and the problematic film on the tip wall to be discarded. This method is more accurate because the dispensed volume is not affected by the liquid film retained in the tip. Common errors to avoid include pipetting too rapidly, which creates inconsistent aspiration, and holding the pipette horizontally, which can cause liquid to seep into the internal mechanism.
Ensuring Precision and Longevity
Micropipettes are precision instruments that require regular care to maintain accuracy over time. The most important maintenance practice is regular calibration, typically recommended every three to six months depending on usage frequency. The industry standard for checking performance is the gravimetric method, which involves weighing the mass of distilled water dispensed. This mass is then converted to volume using the known density of water, allowing technicians to verify the pipette’s accuracy against its set volume.
Routine cleaning and decontamination are important to ensure the longevity and safety of the instrument. If the pipette is used with biohazardous or corrosive materials, it must be properly cleaned to prevent damage to internal components and eliminate cross-contamination. When not in use, proper storage involves placing the micropipette vertically in a dedicated stand or carousel. Storing the instrument upright prevents residual liquid from flowing into the internal mechanism and relieves tension on the internal spring if the volume is set to its maximum.