Advanced Micropipetting Techniques (2024) for Lab Work

Need to nail down the art of micropipetting? From basics to choosing the right type to tricks for perfect precision, this article breaks down everything you need to know to carry out accurate micropipetting that supports dependable lab results.

Key Takeaways

Micropipettes are crucial for precise liquid handling in labs, with different types catering to various liquids: air displacement for aqueous solutions, positive displacement for viscous/dense liquids, and multi-channel for high-throughput operations.
Proper micropipetting depends on a strong understanding of the device’s components, techniques, and the consistent application of good pipetting practices, including handling the plunger correctly and avoiding contamination.
Regular maintenance and calibration of micropipettes are essential for accuracy and precision, necessitating careful volume adjustment, tip selection, and consideration of environmental factors.

Understanding Micropipettes: Types and Functions

The micropipette, a staple in any laboratory setting, is a device used for the precise measurement and transfer of small volumes of liquid. Its operation involves a plunger mechanism and a disposable tip, used for aspirating and dispensing liquids. Moreover, the mastery of micropipette techniques is crucial for laboratory professionals to ensure accurate and consistent results.

Furthermore, There are three main types of micropipettes, each with its specific uses and functions based on the nature of the liquid being handled – air displacement micropipettes, positive displacement micropipettes, and multi-channel micropipettes.

Air Displacement Micropipettes

Air displacement micropipettes are particularly useful for handling aqueous and non-viscous liquids. They operate on the principle of air displacement, where a micropipette plunger is used to draw in or dispense liquid from the pipette tip. This method is preferred for its high precision and accuracy, particularly when dealing with very small volumes of liquid.

Positive Displacement Micropipettes

On the other hand, positive displacement micropipettes come into play when dealing with challenging liquids such as viscous, volatile, or dense substances. This type of micropipette involves a piston directly contacting and displacing the sample, eliminating the need for an air cushion and reducing thermal disequilibrium errors.

These micropipettes are particularly effective for handling high-density solutions, foaming liquids, substances sensitive to temperature or pressure changes, and organic solvents.

Multi-Channel Micropipettes

When it comes to high-throughput laboratory operations, multi-channel micropipettes enhance efficiency and versatility by processing multiple samples simultaneously. They are integral in performing specialized tasks such as:

ELISA tests
Molecular screenings
Tissue culture
Drug screening
DNA amplification

With various configurations available, including 8-channel, 12-channel, and 16-channel variants, multi-channel micropipettes offer flexibility to meet different laboratory requirements and increase productivity in laboratory settings.

Micropipette Components and Proper Handling

A fundamental understanding of the basic components of a micropipette is essential for its proper handling and operation. The components of the device include:

The plunger
The digital display
The tip cone
The tip ejector
The grippy handle

Each of these parts plays a crucial role in the overall functionality of the device. The digital display indicates the volume of the liquid being aspirated or dispensed, which can be adjusted by rotating the plunger.

Careful manual handling of the micropipette, such as avoiding contact with the micropipette tips and using racks to prevent damage and contamination, is necessary to ensure accurate and precise measurements.

Plunger and First/Second Stops

The plunger of the micropipette is a key component that controls the aspiration and dispensing of the liquid. It is designed with two stops: the first stop is used to aspirate the tip with the selected volume of liquid, and the second stop is used to dispense the tip’s contents completely, including the ‘last drop’.

It’s important to operate the pipette plunger slowly and steadily, especially when handling viscous liquids, to ensure accurate and precise measurements.

Digital Display and Volume Adjustment

The digital display on a micropipette shows the volume of liquids you will aspirate or dispense, often using color coding for different volume units. You can adjust the volume using the volume adjustment knob until the correct volume appears on the micropipette.

However, take care not to exceed the micropipette’s maximum capacity or force the volume adjustment knob, as this may damage the device.

Tip Cone and Ejector

The tip cone and ejector are another two critical components of a micropipette. The tip cone ensures a secure fit for standard pipette tips, which is vital for achieving accurate volume measurements. The ejector button, on the other hand, is designed for efficient tip release post-pipetting, reducing potential contamination risks.

Thus, using filter tips is recommended to safeguard the micropipette against contaminants, thereby reducing the frequency of corrective maintenance required.

Essential Pipetting Techniques

There are two essential pipetting techniques that laboratory professionals must master to ensure accurate and consistent results: forward pipetting and reverse pipetting. Forward pipetting is the process commonly used for transferring aqueous solutions and involves a two-step plunger action.

Reverse pipetting, on the other hand, is suitable for viscous or volatile liquids and leaves residual liquid in the tip to ensure complete dispensing of the precise volume. Consistent training in these pipetting techniques is crucial for achieving precision and accuracy in laboratory results.

Forward Pipetting

The forward pipetting technique involves the following steps:

Press the operating button of the micropipette to the first stop to aspirate the tip with the selected volume of liquid.
Submerge the tip into the solution to a depth of 1 cm.
Slowly release the operating button to draw in the liquid.

To dispense the liquid, the operating button is gently pressed down to the second stop while inside the receiving vessel. After dispensing, the tip is withdrawn from the vessel while sliding the thumb along the vessel’s side to promote full emptying, completing the pipetting cycle.

Reverse Pipetting

The reverse pipetting technique, on the other hand, is particularly beneficial for scientists working with substances that are viscous, produce foam, or have high vapor pressure. This technique involves an over-extension of the plunger to the second stop, also known as the blowout stop, before the micropipette tip is submerged in the liquid.

The plunger is then slowly released back to its full extension to aspirate and eject the liquid. The liquid is dispensed by:

Pressing the plunger to the first stop
Maintaining a small volume within the tip
Expelling the liquid into a separate container to ensure the volume dispensed is accurate.

Achieving Accuracy and Precision in Micropipetting

Achieving high levels of accuracy and precision in micropipetting is essential for reliable laboratory results. Some of the ways to ensure this include maintaining the correct immersion angle and depth, pre-rinsing pipette tips before use, and controlling the speed and rhythm of pipetting.

Moreover, firmly attaching the pipette tip onto the tip cone is crucial to prevent air leaks that can negatively impact accuracy.

Immersion Angle and Depth

The immersion angle and depth play a vital role in accurate liquid aspiration and dispensing. When pipetting, it is recommended to hold the pipette vertically, especially for volumes under 50 μl. The tip of the pipette should be immersed 2-3 mm into the liquid when aspirating to avoid drawing in bubbles and excess liquid.

The immersion depth varies with the pipette type, with micro-volume pipettes requiring an immersion depth of 1-2 mm, and normal range pipettes requiring an immersion depth of up to 3-6 mm.

Pre-rinsing, Speed, and Rhythm

Pre-rinsing pipette tips before use can enhance accuracy by neutralizing the capillary effects and adjusting the air temperature inside the tip to match the sample. The speed at which the liquid is aspirated and dispensed should also be controlled and steady to enhance the reliability of the micropipetting technique.

Moreover, practicing and maintaining a consistent rhythm during pipetting lab sessions contribute significantly to the reproducibility and accuracy of laboratory results.

Environmental Factors Affecting Micropipetting

Environmental factors such as temperature and humidity can influence micropipetting accuracy and precision by altering the physical properties of the liquids and the internal mechanics of the micropipette. For instance, sudden temperature changes might reduce the accuracy of micropipetting due to the expansion or contraction of liquid volumes and air spaces within the micropipette.

Therefore, regular calibration checks under specific laboratory conditions are necessary to ensure accurate pipetting outcomes.

Temperature and Humidity

Temperature and humidity can have significant effects on micropipetting accuracy. For instance, micropipettes can over-deliver cold liquids and under-deliver warm liquids, with inaccuracies as high as 37% due to thermal disequilibrium. Moreover, the potential for evaporation, which can lead to pipetting inaccuracies, is considerably reduced in high humidity conditions.

Therefore, to reduce temperature-related inaccuracies in micropipetting, it is advised to equilibrate the fluids and the liquid handling device to the same temperature in the same environmental conditions.

User Habits and Ergonomics

User habits and ergonomic considerations can significantly affect pipetting tasks. A well-designed ergonomic micropipette should have a comfortable grip, be lightweight, and be without sharp edges to minimize contact stress and strain during pipetting tasks. Regular breaks and varied work routines are also recommended, especially for tasks that involve repetitive movements.

While electronic pipettes reduce the force required for operation, they may cause strain due to their additional weight during extended use.

Micropipette Maintenance and Calibration

Regular maintenance of micropipettes is vital to ensure precision and consistency in measurements. This is particularly crucial after the micropipette is cleaned or when it is used with reactive chemicals.

Moreover, calibration, which involves measuring the accuracy of the micropipette by dispensing water into a clean beaker and repeating the measurement several times, is important to verify that the micropipette is functioning properly.

Maintenance Tips

Proper micropipette care and handling involve several maintenance practices. These include:

Avoiding over-pressing the plunger beyond the designed stops
Not turning the volume adjustment knob past the micropipette’s volume limits
Ejecting tips into an appropriate waste container to reduce contamination.

Store micropipettes in a clean, cool, and dry area, adhering to recommended temperature and humidity levels. Regularly maintain and calibrate micropipettes, especially when used with reactive chemicals, to ensure consistent accuracy and precision.

Calibration Procedures

Calibration is an important procedure that verifies the accuracy of micropipettes. Therefore, it must comply with the ISO 8655 standard, which specifies requirements for the calibration of piston-operated volumetric equipment, and the ISO 17025 standard, which sets the competency standard for calibration labs.

Professionals or students should calibrate single and multi-channel micropipettes every 3 to 6 months. Test at least two volumes, both nominal and lowest settings, with 10 replicates for each. Verify the calibrated volume by weighing the dispensed water in a clean beaker using a balance. Follow detailed tutorials to ensure accuracy.

Micropipetting Applications and Benefits

Micropipettes are indispensable tools in various fields and industries due to their ability to:

Measure and transfer small volumes of liquid with high precision and accuracy
Increase the efficiency and performance of laboratory testing
Contribute to more reliable results.

Whether in experimental or diagnostic laboratories, micropipettes are considered a reliable tool for precise liquid handling.

Fields and Applications

Micropipettes are commonly used in various fields, including:

Cosmetic production
DNA sampling
PCR workflows
Molecular biology
Forensic science
Environmental research
Food and beverage manufacturing
Sample preparation in high-performance liquid chromatography (HPLC)

In life sciences applications, such as molecular biology research, DNA RNA purification, PCR, RT PCR, cloning, proteomics, and drug discovery, micropipettes are essential for precision in experiments.

The pharmaceutical, healthcare, and food and beverage industries, as well as environmental monitoring and academic and research institutes, also rely on micropipettes for precise liquid handling in the production of diagnostic kits, and in research and quality control, including analytical chemistry applications.

Benefits of Micropipetting

The benefits of micropipetting are numerous. They offer high levels of accuracy and precision in liquid handling, which is essential for the reliability of laboratory test results and adherence to Good Laboratory Practices (GLPs). The convenience of micropipette starter kits and their efficient design enables quick and efficient transfer of precise volumes of liquid, thus saving time in the laboratory.

Moreover, micropipettes are designed for easy operation, with simple volume adjustments contributing to their user-friendliness. Micropipetting is, therefore, a fundamental skill in research labs, particularly in fields like pharmaceuticals, where tasks such as sampling and analysis of various substances are critical to experiments and studies.

Tips for Choosing the Right Micropipette

Choosing the right micropipette for your specific needs can be a daunting task. However, by considering factors such as:

the specific minimum and maximum volume range
universal tip cone
autoclavable nature
handling comfort

You can ensure that you select a micropipette that is not only accurate and precise but also ergonomic and user-friendly.

It’s also important to purchase micropipettes from reputable brands to ensure long-term reliability and proper support for maintenance and calibration.

Volume Range

When selecting a micropipette, the specific minimum and maximum volume range is a critical factor to consider. Using larger pipettes to dispense smaller volumes can lead to decreased accuracy, especially within the critical 1-10 µL volume range. Therefore, for the most precise measurements, selecting the smallest micropipette capable of dispensing the desired volume is recommended.

Moreover, matching the to use a micropipette upto the volume of liquid needed can increase accuracy over using a pipette with a broader volume range.

Ergonomics and Quality Standards

Ergonomics and quality standards are another two important factors to consider when choosing a micropipette. A well-designed ergonomic micropipette should have a comfortable grip, be lightweight, and be without sharp edges to minimize contact stress and strain during pipetting tasks.

In terms of quality standards, using appropriate tips that allow for clear visual quality checks is recommended. Colored pipette tips are generally discouraged as they may conceal product imperfections, potentially compromising the results.

Summary

In conclusion, mastering the art of micropipetting is a crucial skill for laboratory professionals. From understanding the different types of micropipettes and their functions to learning the essential pipetting techniques, maintaining the pipettes, and choosing the right device, each step of adjustable micropipette plays a vital role in ensuring accurate and precise measurements in laboratory practices. Regardless of the field or application, micropipettes prove to be an indispensable tool in any laboratory setting, offering numerous benefits and contributing to more reliable and reproducible results.

Certified MTP has an elite selection of lab solutions, Measuring Pipette, Mohr Type Measuring Pipettes, showcasing industry-leading brands including Flasks, Beakers, Pipettes, & Graduated Cylinders, Erlenmeyer Flasks, Measuring Pipettes, Stainless Steel Beakers, Glass Graduated Cylinders, and Plastic Graduated Cylinders.

Frequently Asked Questions

What is micropipetting technique?

Micropipetting technique involves gradually drawing up the plunger to avoid drawing in air, and then pressing the plunger to release the reagent into the destination tube. Remember to utilize the second stop to expel the full volume from the tube to the tip.

What is the use of a micropipette?

A micropipette is used in the laboratory to accurately measure and transfer small amounts of liquids, typically between 1 and 1000 microliters. It’s a precise tool for transferring fluids.

What is the aim of micropipetting?

The aim of micropipetting is to accurately measure and dispense very small volumes of liquid, typically in molecular biology and chemistry labs. It allows for precise control over the amount of liquid being transferred.

What is the difference between pipetting and micropipetting?

The main difference between pipetting and micropipetting is the volume they can accurately measure and deliver. Pipettes handle larger volumes, typically starting at 1 milliliter, while micropipettes measure between 1 and 1000 microliters.

What are the different types of micropipettes?

There are three main types of micropipettes: air displacement micropipettes, positive displacement micropipettes, and multi-channel micropipettes. The choice of type depends on the properties of the liquid being handled.