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Micropipettes – Parts, Types, Applications, Techniques

Micropipettes – Parts, Types, Applications, Techniques

Table of Contents

What is a micropipette?

  • A micropipette is a laboratory instrument designed specifically for precise and accurate measurement and transfer of small volumes of liquid samples, typically in the microliter range (µl). While regular pipettes or glass pipettes are suitable for transferring larger volumes in milliliters, micropipettes are essential tools for laboratories that deal with small sample sizes.
  • Micropipettes find applications in a wide range of scientific fields, including pharmaceutical research, molecular biology, forensic analysis, and diagnostic laboratories. They enable researchers and technicians to handle minute quantities of liquids with high precision and reproducibility, which is crucial for conducting experiments and tests.
  • There are different types of micropipettes available, categorized based on various factors such as their operating principle, capacity, channels, and working mechanism. Regarding the operating principle, micropipettes can be classified into air displacement and positive displacement pipettes. Air displacement pipettes work by creating a vacuum to aspirate the liquid into the pipette tip, while positive displacement pipettes use a disposable piston to physically displace the liquid.
  • Micropipettes also come in a range of capacities, from P20 (20 µl) to P10,000 (10,000 µl or 10 ml), allowing users to select the appropriate pipette size for their specific needs. Additionally, micropipettes can have a single channel or multiple channels, with the latter enabling simultaneous transfer of multiple samples. The working mechanics of micropipettes can be either manual or automatic, with some models incorporating electronic features for enhanced functionality and ease of use.
  • The development of the micropipette was a significant advancement in the field of liquid handling. Prior to its invention, liquid transfer methods existed, but accurately handling small volumes was challenging. In 1957, Heinrich Schnitger, a Postdoc at the University of Marburg in Germany, invented the first micropipette using a fragile glass capillary tube. However, it required expert handling and was not practical for widespread use.
  • To overcome these limitations, Dr. Warren Gilson introduced the piston-based adjustable micropipette, commonly known as the Gilson Pipetman, in 1960.
  • This innovation revolutionized precise liquid handling by allowing users to control the aspiration and dispensing of small liquid volumes. The Gilson Pipetman served as the foundation for subsequent advancements in micropipette technology.
  • Since then, various companies have further refined and improved micropipettes, introducing new features and enhancing their overall performance. Today, micropipettes are indispensable tools in laboratories worldwide, enabling researchers and technicians to accurately and efficiently handle small volumes of liquids for various applications, contributing to the advancement of scientific knowledge and discoveries.
Components of a micropipette
Components of a micropipette

What is the purpose of pipetting?

Pipetting is the process of accurately measuring and dispensing specific volumes of liquid using a laboratory tool called a pipette. The main purpose of pipetting is to ensure precise and accurate manipulation of small volumes of liquid, which is essential in various laboratory applications such as molecular biology, biochemistry, genetics, and cell culture.

In a laboratory setting, pipetting serves several important functions:

  1. Preparing solutions and reagents: Pipettes are commonly used to measure and dispense precise volumes of liquid to create solutions or prepare reagents for experiments. By accurately measuring the desired volume, scientists can ensure the correct concentrations of components in their solutions.
  2. Diluting samples: Pipettes are employed to accurately measure a specific volume of a sample and then add an appropriate volume of solvent to create a dilute solution. Dilutions are frequently required in analytical procedures, allowing for better analysis within the desired concentration range.
  3. Mixing samples: Pipettes facilitate the transfer of small volumes of liquid from one container to another, enabling scientists to mix samples before analysis. This is particularly useful when combining different reagents or when ensuring homogeneity of a sample.
  4. Transferring samples: Pipettes are used to transfer small volumes of liquid from one container to another. This is important for various tasks, such as preparing samples for analysis or transferring samples to storage containers.
  5. Measuring and dispensing small volumes: Pipettes excel at accurately measuring and dispensing small volumes of liquid, making them indispensable in techniques like polymerase chain reaction (PCR), enzyme-linked immunosorbent assay (ELISA), and other types of assays. These assays often require precise and precise addition of reagents in microliter or sub-microliter quantities.

Overall, pipetting plays a crucial role in laboratory applications by allowing scientists to handle small volumes of liquid with precision and accuracy. It enables the creation of specific solutions and reagents, facilitates the measurement and transfer of samples, and ensures reliable and accurate results in experiments. By mastering pipetting techniques and using quality pipettes, scientists can perform their tasks with greater confidence and achieve optimal outcomes in their research and analysis.

How to Use a Micropipette?

To effectively use a micropipette and ensure its proper functioning and longevity, follow these steps:

  1. Select the appropriate micropipette: Different micropipettes have fixed volume ranges. For example, a P1000 micropipette measures volumes from 100 to 1000 µl, while a P20 micropipette aspirates fluids from 2 to 20 µl. Choose the micropipette that matches the volume of liquid you need to handle.
  2. Adjust the micropipette to the desired volume: After selecting the correct micropipette, use the volume adjustment knob to set the desired volume. Avoid turning the knob beyond the micropipette’s specified range, as this can damage the instrument.
  3. Attach the pipette tip: Select an appropriate pipette tip based on the intended use of the micropipette. Avoid touching the tip with your hands. Instead, place the micropipette above the tip and press it down gently until the tip attaches securely to the tip cone.
  4. Aspirate the liquid: The plunger of the micropipette typically has two stops. To aspirate the liquid, press the plunger down to the first stop. Then, immerse the tip into the liquid you want to aspirate. Release the plunger slowly, ensuring the tip remains inside the liquid until the plunger returns to its original position.
  5. Dispense the liquid: Insert the tip of the micropipette into the container where you intend to dispense the sample. Press the plunger down to the first stop and briefly pause. Then, press the plunger down to the second stop to completely expel the liquid from the tip. Hold the plunger in this position until you have fully removed the tip from the container to prevent any re-aspiration.
  6. Remove the tip: Position the micropipette above a disposal container. Press the tip ejector button or lever to safely and hygienically remove the used tip. Avoid using your hands to remove the tip, as this can result in contamination.

By following these steps, you can effectively and accurately use a micropipette, ensuring precise liquid handling and maintaining the instrument’s performance and longevity. Remember to handle the micropipette with care and follow any additional instructions provided by the manufacturer.

Parts of a micropipette

Components of a micropipette
Components of a micropipette

Micropipettes consist of several essential parts that enable precise and accurate liquid handling. Here are the main parts of a micropipette:

  1. Plunger: The plunger is the topmost part of the micropipette. It is used to adjust the volume, aspirate, and dispense the required amount of the sample. By rotating the plunger clockwise or counterclockwise, the volume can be increased or decreased. It typically has two stops, allowing for forward and reverse pipetting.
  2. Tip Ejector: The tip ejector is a mechanism that helps in the easy removal of micropipette tips. It is usually located below the plunger, and by pressing the ejector, the tips can be safely released without the need to touch them.
  3. Volume Window: The volume window is a transparent area on the micropipette body that displays the adjusted volume. It allows the user to visually confirm the selected volume before pipetting.
  4. Shaft: The shaft is a tube-like structure inside the micropipette that is filled with air. It plays a crucial role in the pipetting mechanism by pushing or pulling the liquid as the plunger is operated.
  5. Micropipette Tips: Micropipette tips are disposable attachments that are placed on the end of the micropipette. They come in various sizes to accommodate different sample volumes and are made of materials such as virgin polypropylene or molded plastics. The tips directly come in contact with the liquid being pipetted and are essential for accurate and contamination-free transfer.
  6. Digital Volume Display Window: This feature is present in electronic micropipettes. It displays the volume that the micropipette can withdraw or dispense. The digital display provides precise and clear volume readings, enhancing the convenience and accuracy of pipetting.
  7. Plastic Shaft: The plastic shaft is a tube-like structure filled with air in air displacement micropipettes. It aids in the displacement of air and liquid during the pipetting process. When the plunger is pressed, air from the shaft is released, facilitating liquid aspiration. Releasing the plunger allows the shaft to refill with air, assisting in liquid dispensing.
  8. Ejector Arm: The ejector arm is an extension of the tip ejector button. When the ejector button is pressed, the ejector arm applies pressure on the pipette cone, aiding in the secure ejection of used pipette tips.
  9. Tip Cone: The tip cone is the part of the micropipette where the pipette tip is attached. It ensures a secure and tight connection between the micropipette and the tip, preventing any leaks or sample loss during pipetting.
  10. Pipette Tip: The pipette tip is an essential component that comes in direct contact with the liquid. It is made of high-quality materials and designed to be compatible with specific micropipette models. Pipette tips are available in various sizes and can accommodate different volumes of liquid.

Micropipette Tips

Micropipette tips are essential components of micropipettes and are available in various colors and sizes. Here is some information about micropipette tips based on the content provided:

  1. White Micropipette Tips:
    • White micropipette tips are commonly used with P2, P10, and P20 micropipettes.
    • Micro white tips: These tips are designed for P2 and P10 micropipettes, allowing measurement of liquid volumes ranging from 0.2 µl to 10 µl.
    • Medium white tips: Medium white micropipette tips are used with P20 micropipettes and can handle liquid volumes ranging from 2 µl to 20 µl.
  2. Yellow Micropipette Tips:
    • Yellow micropipette tips are specifically designed for P100 and P200 micropipettes.
    • They are used for measuring liquid volumes between 20 µl and 100 µl.
  3. Blue Micropipette Tips:
    • Blue micropipette tips are utilized with P1000 micropipettes, which are capable of handling higher liquid volumes.
    • These tips can measure liquid volumes from 200 µl up to 1000 µl, which includes milliliter ranges as well.

It’s important to note that the colors mentioned for micropipette tips may vary depending on the manufacturer or supplier. These color codes are typically standardized to help It’s important to note that the colors mentioned for micropipette tips may vary depending on the manufacturer or supplier. These color codes are typically standardized to help users identify the appropriate tip size and compatibility with specific micropipettes. Using the correct micropipette tip size ensures accurate and reliable liquid handling.

Attaching Micropipette Tip – How to Attach a Micropipette Tip?

Attaching a micropipette tip is a crucial step in ensuring accurate and contamination-free liquid handling. Here is a procedure to properly attach a micropipette tip:

  1. Select the Proper Sized Tip:
    • Choose a micropipette tip that is compatible with the volume range you intend to pipette.
    • Ensure that the tip size matches the specific micropipette model you are using.
  2. Open the Box of Micropipette Tips:
    • Open the box of micropipette tips carefully, avoiding any contact with the tips to maintain sterility.
  3. Attach the Micropipette Tip:
    • Hold the micropipette by the barrel, away from the tip end.
    • Align the open end of the tip with the nozzle or shaft of the micropipette.
    • Insert the micropipette shaft into the tip opening until it is fully seated.
    • Apply gentle downward pressure to firmly attach the tip to the micropipette.
    • Ensure that the tip is securely attached without any wobbling or looseness.
  4. Remove the Micropipette with Attached Tip:
    • Carefully lift the micropipette, making sure the attached tip remains in place.
    • Avoid any contact between the tip and surfaces that may introduce contamination.
  5. Close the Box of Micropipette Tips:
    • Close the lid or cover of the box of micropipette tips without touching the tips directly.
    • This helps maintain the sterility and cleanliness of the remaining tips for future use.

Proper attachment of the micropipette tip is essential to prevent leakage, ensure accurate volume delivery, and minimize the risk of sample contamination. Following this procedure helps maintain the integrity and performance of the micropipette during liquid handling tasks.

Types of micropipette

Micropipettes are essential laboratory tools used for accurate and precise measurement and transfer of liquid samples. They come in various types based on different features. Here are the different types of micropipettes:

I) Based on Volume:

  1. Fixed Volume Micropipette: This type of micropipette has a fixed volume capacity for aspirating or dispensing liquid. It is suitable for tasks that require consistent dispensing of the same volume multiple times.
  2. Variable Volume Micropipette: In contrast to fixed volume micropipettes, variable volume micropipettes allow the adjustment of the aspirated or dispensed volume according to specific requirements. They have a range and limit of volume that can be adjusted as needed.

II) Based on Principle of Operation:

  1. Air Displacement Micropipette: Air displacement micropipettes operate by creating a vacuum through the movement of a piston. The set volume is aspirated by immersing the tip in the liquid, and then the liquid is dispensed by pressing the plunger. It is commonly used for standard pipetting applications and offers high accuracy. There are several types of air displacement pipettes available, each offering specific features and capabilities. Here are the different types of air displacement pipettes:
    • Volume Handled Micropipettes: These pipettes are categorized based on the range of volumes they can handle. They include micropipettes designed for handling specific volume ranges, such as microliter (µL) or milliliter (mL) volumes.
    • Adjustable or Fixed Micropipettes: Adjustable micropipettes allow users to set and adjust the desired volume within a specified range. They provide flexibility in pipetting various volumes. In contrast, fixed micropipettes have a fixed volume capacity, suitable for tasks that require consistent volume dispensing.
    • Single-Channel, Multi-Channel, or Repeater Micropipettes: Single-channel pipettes are designed to handle one sample at a time, whereas multi-channel pipettes have multiple channels for simultaneous aspiration and dispensing of several samples. Repeater pipettes are specifically designed for repetitive dispensing of the same volume.
    • Manual or Electronic Micropipettes: Manual micropipettes require manual operation by the user, typically using a plunger or button for aspiration and dispensing. Electronic micropipettes, on the other hand, are automated and feature electronic controls for precise volume adjustment and pipetting.
    • Cylindrical or Conical Tips Micropipettes: The tips of air displacement pipettes can be cylindrical or conical in shape. Cylindrical tips are straight and have a uniform diameter throughout, while conical tips taper towards the end. The choice of tip shape depends on the specific application and sample type being pipetted.
    • Locking or Standard Micropipettes: Locking micropipettes have a locking mechanism that secures the volume setting, preventing accidental changes during pipetting. Standard micropipettes do not have a locking feature and require careful handling to maintain the set volume.
  2. Positive Displacement Micropipette: Positive displacement micropipettes function by moving the piston down into the tip to aspirate the liquid and then moving it up to dispense the set volume. This type of micropipette is suitable for viscous or volatile liquids that may stick to the tip.

III) Based on Operating Mechanism:

  1. Mechanical Micropipette: Mechanical micropipettes operate using a spring piston mechanism and are manually operated by hand. They are widely used and require the user to depress the plunger for aspiration and dispensing.
  2. Electronic Micropipette: Electronic micropipettes are automatic and feature a touch-button operation. They enable the precise aspiration and dispensing of the sample with a single touch. They are suitable for applications that require high accuracy and reproducibility.

IV) Based on Number of Channels:

  1. Single-Channel Micropipette: Single-channel micropipettes have a single channel for aspirating and dispensing samples. They are commonly used for routine pipetting tasks.
  2. Multi-Channel Micropipette: Multi-channel micropipettes have multiple channels, usually 8, 12, or 16, allowing simultaneous aspiration and dispensing of multiple samples. They are designed to reduce workload and increase efficiency in high-throughput applications.

These different types of micropipettes provide researchers with options to suit their specific needs for accurate liquid handling and sample manipulation in various laboratory settings.

Working mechanism of Micropipette

The working mechanism of a micropipette involves the precise manipulation of air displacement and the movement of liquid through the pipette tip. Here’s an explanation of how a micropipette works:

  1. Aspiration: When the plunger of the micropipette is pressed down, it creates a partial vacuum inside the shaft, typically filled with air. This decrease in pressure causes the air inside the shaft to expand, pushing against the liquid in the micropipette tip. As a result, the liquid is drawn into the tip due to the difference in pressure between the tip and the sample.
  2. Liquid Collection: As the plunger is released, it moves upward, reducing the volume inside the shaft. This upward movement of the piston creates a positive pressure inside the shaft, causing the air to be replaced by the liquid present in the micropipette tip. The liquid is securely held in the tip due to the cohesive forces between the liquid molecules and the hydrophobic properties of the tip material.
  3. Dispensation: To dispense the collected liquid, the micropipette tip is carefully placed into the desired container or vessel. By gently pressing the plunger down to the first stop, a small amount of liquid is expelled from the tip. This partial dispensation allows for precise control over the amount of liquid released.
  4. Complete Dispensation: To completely dispense the remaining liquid, the plunger is pressed further down to the second stop. This action forces the piston to move upwards again, expelling the remaining liquid from the micropipette tip.

Throughout the pipetting process, the volume adjustment knob on the micropipette allows users to set the desired volume of liquid to be aspirated and dispensed. By rotating the knob, the distance traveled by the plunger inside the shaft can be adjusted, thereby determining the volume of liquid transferred.

It is important to note that accurate pipetting requires careful technique and attention to details such as ensuring the tip is fully submerged in the liquid during aspiration and avoiding air bubbles. Proper maintenance, regular calibration, and using appropriate pipetting techniques are crucial to ensure precise and reproducible results when using a micropipette.

Volume adjustment on Micropipette – How to Adjust Volume in Micropipette?

The volume adjustment on a micropipette is a crucial feature that allows users to set the desired volume for accurate liquid measurement. Here is some information about volume adjustment on a micropipette based on the provided content:

  1. Adjusting the Volume:
    • To adjust the volume, locate the volume adjustment dial situated at the top of the micropipette.
    • Rotate the dial either to the right or left to decrease or increase the volume, respectively.
    • As you rotate the dial, the digital layout on the micropipette displays the corresponding volume.
    • The digital readout shows the volume that has been set for measurement.
  2. Digital Readout on Micropipette:
    • The digital readout on the micropipette consists of three numbers, representing different volume ranges for various sizes of micropipettes.
    • The numbers on the digital readout correspond to the volumes set on different sizes of micropipettes.
  3. Example of Volume Representation:
    • P1000 Micropipette: The bottom number represents 10’s of microliters (µl), the middle number represents 100’s of µl, and the top number represents 1000’s of µl.
    • P200 Micropipette: The bottom number represents 1’s of µl, the middle number represents 10’s of µl, and the top number represents 100’s of µl.
    • P20 Micropipette: The bottom number represents 0.1 of µl, the middle number represents 1’s of µl, and the top number represents 10’s of µl.

These volume representations may vary slightly depending on the specific micropipette model or manufacturer. It is important to refer to the user manual or instructions provided with the micropipette to understand the volume adjustment settings specific to your micropipette model.

Size and Range / Technical Specifications of Micropipette

Micropipettes are offered in a variety of sizes ranging from 0.1 up to 10,0000 ul. The most commonly used versions of micropipettes that have a single channel of variable volume are listed below with their error tolerances as defined in the ISO-8655-2 standards.

They may be referred to as P10 or P20, P1000 or P5000 pipettes based upon the largest volume of liquid that is aspirated and disseminated by the pipette. For example the 0.5-10ul micropipette is often called P10 pipette.

Volume Range
(ul)
ClassificationIncrement
(ul)
Accuracy
(± %)
Accuracy
(±ul)
CV
(± %)
CV
( ± ul)
0.2-2P20.00220.041.20.024
0.5-10P100.0210.10.50.05
2-20P200.020.80.160.40.08
5-50P500.10.80.40.40.2
10-100P1000.20.60.60.20.2
20-200P2000.20.61.20.20.4
100-1000P10001.00.660.22
500-5000P500010.00.6300.210
1000-10000P1000020.00.6600.220

The measuring ranges for the micropipettes most frequently used are:

  • P10 – 1.0-10.0µL – White tips (slightly translucent)
  • P20 – 2.0-20.0 µL – Yellow tips
  • P100 -10.0-100.0 µL – Yellow tips
  • P200 – 20.0-200.0 µL – Yellow tips
  • P1000 – 11000.0 µL –Blue tips (should only be used for volumes 200 or greater)

Working Principle of micropipette

Air displacement micropipettes function via the piston driven air displacement. As the piston pulled down, air contained within the sleeve of the micropipette is released because of the force with which the liquid in the micropipettes ‘ tip is also eliminated.

As the piston is moved upwards there is a vacuum created in the empty space left due to the movement of the piston. The air at on the tips to expand to fill in the empty space. The tip air is replaced by the liquid that is drawn upwards to the tip.

Positive displacement micropipettes work via piston-driven displacement. The piston inside the positive displacement micropipette is directly in touch with liquid. If the piston gets pulled down, liquid that is inside the sleeve of the micropipette is also moved downwards and is ejected from the tips. As the piston moves upwards it draws the liquid with it in an upward direction.

Using an Air Displacement Micropipette

Accurate measurement of liquids is dependent on the proper micropipette use. Air displacement micropipettes operate using the principle of air displacement. The plunger is compressed by the thumb, and when it releases the liquid is drawn towards the tip of a disposable. If the plunger is then pressed again and the liquid is dispersed. Between these stages, there are many smaller steps that assist in making the process of liquid dispensing more precise.

Using an Air Displacement Micropipette
Using an Air Displacement Micropipette | Image Source: https://www.microlit.us/micropipette-product-guide/

Position 1

  • In this, the micropipette is at rest position: You can press-fit the tip of the micropipette, but not touching directly to the edge.

Position 2

  • In this, the plunger is depressed till the first stop: To aspirate the liquid at the tip push the plunger until its first stop. Place the pipette tip horizontally within the liquid.
  • Release the plunger: Release the plunger slowly while the tip is submerged. The liquid will aspirate into the tip of the pipette. The liquid is then poured into the tip according to the micropipette’s volume that is set.

Position 3

  • Depress the plunger: place the tip on the interior wall of the vessel at an angle of steep. The plunger should be pressed slowly to the first stop in order to disperse the liquid. To completely empty the tip push the plunger until next stop. Clean the tip with the wall before taking the tip from the vessel.

Some micropipettes are able to deliver a set amount of liquid. But, the majority of them can be adjusted with their variable volume settings. Variable volume micropipette is available with different ranges of measurement and upper and lower limit of measurement. In these situations, the error percentages can differ as according to the liquid being measured. Dispensing lesser than the lowest limit of the range could result in inaccurate measurements of liquid while attempting to use the upper limit will fill the tip and allow liquid to flow into the pipette’s body.

Cleaning, maintenance and storage of a micropipette

Maintaining and caring for the micropipette is a crucial routine in labs. Following a well-planned maintenance plan will reduce the expense of purchasing the expensive equipment. Cleaning the micropipette is a process that requires time, effort and focus as otherwise, the micropipette could be damaged. It is therefore essential to be careful when cleaning it. This is a complete guide to cleaning micropipettes.

External Cleaning

The majority of the pipette is able to be cleaned using the typical lab cleaning chemicals such as soaps, alcohol or soaps. To ensure complete sterilization you should allow the cleaning solution to sit on the micropipette 10 to 15 minutes before taking it off with a cloth.

Internal Cleaning

Cleaning the inside of the micropipette may be time-consuming as it requires complete disassembly. Additionally, every component of the micropipette must be cleaned in a proper manner based on the fluid used in the sampling.

  • Check the instruction manual for precise instructions for micropipette
  • Make use of a cotton swab soaked in the cleaning solution and distil water.
  • Make sure to lightly grease your pistons using the lubricant supplied upon purchase.
  • Assemble all the components and test to make sure the micropipette is operating efficiently.

Contamination Cleaning

If the micropipette is found to be contaminated by a known ingredient, then there’s certain cleaning procedures that need to be followed based on the kind of substance. Cleaning the micropipette with the above procedure won’t suffice if the micropipette is cross-contaminated.

Solution TypesProcess to clean
For aqueous solutions, organic solvents and proteinsRinse the contaminated parts with distilled water or 70 percent ethanol and air dry at approx 60°F temperature.
For infectious liquidsAutoclave the lower section at a temperature of 120°C for 15-20 minutes then allow it to return at room temperature before reassembling.
For radioactive substancesPlace the pipette in a solution like Decon and then rinse and air dry.
For nucleic acidsBoil lower parts of micropipette in glycine/ HCI buffer (pH2) for 10 minutes, rinse with distilled water, and air dry.

A clean and well-maintained pipette will protect the lab from potentially harmful substances. This makes the pipette more efficient, durable, and reliable and also reduces the expense of sample collection.

The proper storage of micropipettes is just as crucial as cleaning and calibrating the devices. The micropipette as well as accessories must be kept in a cool, clean and dry area. The storage space should have temperatures ranging between -20°C and 50° degC (from -4 degF up to 120°F) with the relative humidity ranging between 5 and 95 percent. Another important thing to keep in mind is the fact that your instrument needs to be kept in an upright place. To put pipettes away, Microlit recommends the use of its carousel stand Microlit Faveo.

Different pipetting techniques 

The most frequently employed pipetting techniques comprise forward pipetting and reverse pipetting. Before we learn about these techniques in greater detail the basic guidelines for pipetting that are listed below will be worth mentioning.

  1. The plunger should be released and pressed gradually, always especially when working with viscosity reagents or solutions. Check that the plunger doesn’t snap.
  2. Make sure that the tip is secured to the cone.
  3. Before beginning your experiment, you should fill then empty your tip three times with the reagent , or solution you’ll be pipetting.
  4. Make sure to hold the micropipette upright in a position while you aspirate. The Grippy should rest on your index finger.
  5. Check your tips and the pipette as well as the solution/reagent are all at the same temperature.

Forward Pipetting Technique

The forward pipetting technique is a commonly used method for transferring liquid using a micropipette. Here are the steps involved in the forward pipetting technique based on the provided content:

  1. Adjust the Required Volume:
    • Use the volume adjustment knob on the micropipette to set the desired volume for liquid transfer.
  2. Attach the Suitable Tip:
    • Choose a suitable micropipette tip that matches the volume you have set on the micropipette.
    • Attach the tip securely to the micropipette.
  3. Press the Knob to the First Stop:
    • Gently press the knob of the micropipette to the first stop.
    • This will allow the micropipette to draw up the liquid into the tip.
  4. Dip the Tip into the Solution:
    • Dip the tip of the micropipette into the liquid solution to a certain depth, ensuring it is submerged in the liquid.
  5. Release the Pipette Knob:
    • Slowly release the micropipette knob back to its starting position.
    • This allows the liquid to be drawn into the tip by capillary action.
  6. Withdraw the Tip from the Liquid:
    • Carefully withdraw the tip from the liquid, ensuring that no liquid remains on the outside of the tip.
  7. Move the Pipette to the Receiving Vessel:
    • Position the micropipette tip over the receiving vessel where you want to transfer the liquid.
  8. Dispense the Liquid:
    • Press the micropipette knob to the second stop, which will dispense the liquid from the tip into the receiving vessel.
    • This step also helps to remove any remaining liquid in the tip.
  9. Eject or Release the Tip:
    • After transferring the liquid, use the tip ejector mechanism on the micropipette to safely eject or release the used tip.
    • Dispose of the used tip properly according to the lab’s waste management guidelines.

It is important to follow proper pipetting techniques and practices to ensure accurate and precise liquid transfer while minimizing the risk of contamination or errors.

Forward Pipetting Technique
Forward Pipetting Technique | Image Source: https://www.microlit.us/micropipette-product-guide/

Reverse Pipetting Technique

Reverse pipetting is a technique commonly used when working with volatile liquids or when high precision and accuracy are required. Here are the steps involved in reverse pipetting based on the provided content:

  1. Adjust the Required Volume:
    • Use the volume adjustment knob on the micropipette to set the desired volume for liquid transfer.
  2. Attach the Suitable Tip:
    • Choose a suitable micropipette tip that matches the volume you have set on the micropipette.
    • Attach the tip securely to the micropipette.
  3. Press the Knob to the Second Stop:
    • Instead of stopping at the first stop, press the knob of the micropipette all the way down to the second stop.
    • This prevents the liquid from being drawn into the tip during the aspiration step.
  4. Dip the Tip into the Solution:
    • Dip the tip of the micropipette into the liquid solution to a certain depth, ensuring it is submerged in the liquid.
  5. Release the Pipette Knob:
    • Slowly release the micropipette knob back to its starting position.
    • This allows the liquid to be drawn into the tip by capillary action, but only up to a certain volume.
  6. Withdraw the Tip from the Liquid:
    • Carefully withdraw the tip from the liquid, ensuring that no liquid remains on the outside of the tip.
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  7. Move the Pipette to the Receiving Vessel:
    • Position the micropipette tip over the receiving vessel where you want to transfer the liquid.
  8. Press the Knob to the First Stop:
    • Press the micropipette knob down to the first stop, dispensing the liquid from the tip into the receiving vessel.
    • It is important not to press the knob to the second stop to avoid drawing any residual liquid back into the tip.
  9. Discard or Eject the Tip:
    • After completing the liquid transfer, use the tip ejector mechanism on the micropipette to safely discard or eject the used tip.
    • Dispose of the used tip properly according to the lab’s waste management guidelines.

Reverse pipetting is useful when working with volatile or precious liquids as it minimizes the potential for aerosol formation and ensures more accurate and consistent volume delivery.

Reverse Pipetting Technique
Reverse Pipetting Technique | Image Source: https://www.microlit.us/micropipette-product-guide/

Note: Any residual liquid stays in the end. It is not part of the volume of dispensing.

Choose the right micropipette

Since micropipettes can be used for many different applications The issue is how to select the appropriate micropipette for the specific job.

To do this, follow our easy, step-by-step guide. The first step is writing down the specifics of the experiment you’re conducting using the micropipette. It should include details about the amount of sample to be taken, the volume of the sample to be transferred as well as the number of duplicates and whether you require sterile conditions and other details. In this manner can aid in deciding the best micropipette for you.

Selecting the Type of Micropipette

Micropipettes are generally classified into two kinds namely air Displacement Micropipettes as well as Positive Displacement Micropipettes. It is possible to choose the best type by matching the application to the research you want to conduct.

Next Steps

When you’ve decided on the best type, let’s examine the various sub types that are that are available, based on factors such as the amount of material of work to be done, the quality of tips, the manual or electronic pipettes as well as other factors. This is where the specific details about the procedure will be useful.

1. An adjustable micropipette or a fixed micropipette

You could either choose the micropipette with a volume that is already fixed, or one that is able to be altered as needed. If you typically employ a single volume like 100 ul to conduct your experiments, opt for an instrument with a fixed volume, such as MICROLIT RBO Fixed Volume (Single Channel). If your project requires you working with several volumes, go for the flexible or variable pipette such as MICROLIT RBO Variable Volume (Single Channel or Multichannel).

2. The volume to be handled

As a general rule it is recommended to select the smallest pipette that can handle the required quantity because if the quantity is near the capacity limit of the micropipette accuracy and precision of readings diminish.

  • Single channel or multichannel micropipettes: The choice of this type is according to the quantity of replicates or samples you’re working with. Single channel micropipettes such as those of the MICROLIT RBO Single channel (Fixed volume and variable) are utilized when the quantity of samples is lower, while many sample or even well plate could be handled with ease with multichannel micropipettes such as those of the MICROLIT RBO Multichannel (8-channel and 12-channel).

3. Micropipette Tips

Once you’ve picked the micropipette you want to use then this is the next critical decision you have to make. Micropipette Tips are graduated or not either universal or pipette specific and with filter or no filter, sterile or not, and so on. If the procedure requires aseptic, sterile conditions, you should make use of sterile and filter-filtered tips. Universal tips can be utilized for a range of pipettes. It is crucial to make sure that how much capacity the tip has to that of the micropipette prior making use of.

4. Accuracy with specific tolerance

Micropipettes are made to work with accuracy just a few percent (generally less than 5 percent) of the value you want of the sample. Be sure that your micropipette has the same level of accuracy that you require for the sampling.

Choose the right micropipette
Choose the right micropipette

Micropipette calibration

Micropipette calibration is a crucial element in every laboratory’s procedure to guarantee precise and precise results from pipetting. To ensure that the sample is accurate it is essential to verify the calibration of the pipette each month or after between a few months. Making a correct micropipette calibration as per ISO 8655 standard requires special conditions in the ambient. Things like air pressure or humidity, temperature, or even altitude influence the outcomes of the pipetting. The following text will explain the material or equipment needed and the various steps you should follow and verify the calibration of a micropipette.

Here’s a list things you’ll need to test a micropipette

  • Micropipette that must be calibrated and tips
  • Distilled Water
  • Clean Beaker
  • Thermometer
  • Distilled Water Density Chart and Temperature
  • Semi-Micro Balance having an accuracy of at minimum 0.0001 grams
  • Notebook to write down the results
  • Calculator

How to calibrate the Micropipette?

Calibrating a micropipette is essential to ensure accurate and precise measurements. Here are the steps to calibrate a micropipette:

  1. Prepare the Equipment:
    • Keep distilled water in the refrigerator to ensure a consistent temperature.
    • Use a thermometer to measure the temperature of the distilled water.
    • Clean and dry the beaker on top of the balance.
  2. Check the Micropipette:
    • Ensure that the micropipette is in good condition and functioning properly.
    • Make sure there are no air bubbles in the pipette and that it is clean.
  3. Filling the Micropipette:
    • Fill the micropipette with distilled water, taking care to avoid air bubbles.
  4. Dispensing and Weighing:
    • Dispense the distilled water into the clean beaker on the balance.
    • Use the balance to measure the weight of the dispensed water. Take note of the measurement.
  5. Repeat and Record:
    • Repeat the dispensing and weighing process 5-10 times.
    • Record each measurement accurately for further calculations.
  6. Calculating the Dispensed Volume:
    • Use the formula V = Z * W to calculate the dispensed volume.
    • V represents the amount of dispensed water, W is the weight of the distilled water dispensed, and Z is the conversion factor based on the density of water.
  7. Average the Results:
    • Calculate the average of the dispensed volumes by dividing the total by the number of tests conducted.
  8. Determine the Accuracy:
    • Use the formula A = (100 * Vavg) / V0 to determine the accuracy of the micropipette.
    • A represents the accuracy, Vavg is the calculated volume average, and V0 is the theoretical volume expected to be dispensed.
    • The desired accuracy range is typically between 99% and 101%.
  9. Evaluate the Results:
    • Examine the calculated accuracy results.
    • If the micropipette falls within the acceptable range, it is considered calibrated and safe to use.
    • If the results are inconsistent or outside the acceptable range, the micropipette may need to be adjusted or serviced before further use.

By following these steps, you can ensure that your micropipette is accurately calibrated, providing reliable measurements for your experiments and procedures. Regular calibration and maintenance of micropipettes are important to maintain their accuracy and precision over time.

Sterilization Process of a micropipette

  • The micropipettes are sterilized using autoclaving at temperatures of 121°C (252degF) or 15 PSI for a period of between 15 and 20 minutes.
  • There is no special preparation required.
  • You can utilize steam sterilization bags when you require. When the autoclave is done, the pipette needs to be chilled to ambient temperature for at minimum two hours. Before you pipetting, ensure you have the pipette clean.
  • It is suggested to test the calibration at the end of each sterilization process to get the greatest precision and accuracy.

When is micropipette calibration required?

To maintain the high quality of the results of the sampling The micropipettes need to be in good working order and calibrated properly. Normally, the period that a micropipette has to be calibrated is determined by several factors:

  • Frequency of pipette use
  • The type of liquid that is dispensable via pipettes
  • Care and handling of the pipette
  • Pipette-related applications that require high accuracy also require more frequent calibration

Regulations and standards issued by organizations like The FDA as well as ASTM International provides minimum requirements to ensure the accuracy of laboratory test results. Regulations stipulate that all laboratory equipment used for sampling and production, including micropipettes should be regularly checked at regular intervals.

In the same sequence, the Clinical and Laboratory Standards institute (CSLI) has provided the guidelines for multi-channel and single channel micropipettes. The guidelines state that these micropipettes should be calibrated every 3 to six months. In addition, a minimal of 2 volumes should be tested using 10 copies in both nominal and low settings.

Setting a suitable frequency of calibration minimizes the risk of a wrong liquid delivery in the laboratory . It also ensures transparency, accountability, and confidence in the result.

Uses of Micropipette

Micropipettes are versatile tools used in various laboratory settings due to their precise and accurate measurement capabilities. Here are some common uses of micropipettes:

  1. Clinical and Microbiological Laboratories:
    • Micropipettes are essential in diagnostic tests conducted in clinical and microbiological laboratories. They are used for transferring reagents and samples in serological tests (such as ELISA), molecular tests (PCR, sequencing, blotting techniques), and virus culture in cell lines.
  2. Chemical Laboratories:
    • Micropipettes find applications in chemical laboratories where handling small volumes of viscous and volatile liquids is necessary. They are used in experiments that require precise measurement and dispensing of reagents, solvents, or samples.
  3. Forensic Laboratories:
    • Micropipettes play a crucial role in forensic laboratories during the analysis of blood, tissues, fibers, and DNA materials. They are used to transfer and handle small volumes of samples, aiding in genetic analysis, DNA testing, and the examination of fingerprints found at crime scenes.
  4. Pharmaceutical Laboratories:
    • Micropipettes are utilized in pharmaceutical laboratories for medicine and drug production. They are involved in measuring and dispensing precise volumes of ingredients during the formulation process. Additionally, micropipettes are crucial for quality control (QC) testing of pharmaceutical products to ensure their potency, purity, and uniformity.
  5. Food and Beverage Industry:
    • In the food and beverage industry, micropipettes serve as liquid handling devices. They are employed in tasks such as preparing samples and reagents, diluting solutions, and conducting quality assurance tests to ensure product safety and consistency.
  6. Molecular Biology Laboratories:
    • Micropipettes are extensively used in molecular biology laboratories for a range of experiments. They are vital in techniques like PCR, western blotting, DNA sequencing, and other molecular biology procedures that require accurate measurement and dispensing of small volumes of samples, enzymes, primers, and reagents.

Limitations of Micropipette

Micropipettes, despite their widespread use and importance in laboratories, have certain limitations that should be considered. Here are some common limitations of micropipettes:

  1. Manual Operation and Human Error:
    • Micropipettes require manual operation, and as a result, they are susceptible to human errors. Inconsistent pressure applied to the plunger or misreading the volume markings can lead to inaccurate measurements. Care must be taken to ensure proper technique and minimize the potential for human error.
  2. Cell Damage during Mixing:
    • When mixing cells or sensitive samples, excessive force or vigorous pipetting can cause damage to the cells. The mechanical stress applied during mixing can lead to cell rupture, loss of viability, or altered cellular functions. It is crucial to use appropriate mixing techniques that minimize cell damage.
  3. Potential for Cross-Contamination:
    • If liquid comes into contact with the outside surface of the micropipette tip, cross-contamination can occur. This can happen when the pipette tip touches a contaminated surface or when liquid is accidentally aspirated into the tip cone. Proper pipetting technique and using sterile techniques can help reduce the risk of cross-contamination.
  4. Accuracy with Viscous Liquids:
    • Micropipettes may not provide accurate measurements when handling highly viscous liquids. These liquids tend to stick to the walls of the tip, resulting in incomplete transfer or variations in dispensed volumes. Alternative pipetting methods or specialized pipette tips designed for viscous liquids may be necessary for improved accuracy.
  5. Careful Handling Required:
    • Micropipettes require careful handling to maintain their accuracy and functionality. Dropping or mishandling the micropipette can lead to misalignment, damage to internal components, or calibration issues. Regular maintenance, proper storage, and gentle handling are essential to ensure the micropipette remains accurate and usable.
  6. User-Dependent Accuracy:
    • The accuracy of micropipettes is dependent on the skill and technique of the user. Inconsistent application of pressure on the plunger, improper attachment of the tip, or failure to read the volume markings accurately can result in measurement errors. Proper training, adherence to protocols, and careful attention to detail are necessary to achieve accurate and reliable results.

Despite these limitations, micropipettes remain indispensable tools in many laboratory applications. Recognizing and addressing these limitations through appropriate techniques, protocols, and quality control measures can help mitigate potential errors and ensure the reliable use of micropipettes in scientific research and analysis.

Advantages of micropipette

Micropipettes offer several advantages that contribute to their widespread use in laboratories. Here are some key advantages of micropipettes:

  1. Time Saving:
    • Micropipettes are designed for efficient and precise liquid handling, allowing researchers to quickly and accurately dispense small volumes of liquids. This time-saving feature is particularly valuable when performing repetitive tasks or working with a large number of samples. The precise measurement and dispensing capabilities of micropipettes help streamline laboratory workflows and increase productivity.
  2. Ergonomics:
    • Micropipettes are ergonomically designed to provide comfort and minimize user fatigue during prolonged pipetting tasks. They are lightweight, have a comfortable grip, and feature adjustable plunger forces to accommodate different user preferences. The ergonomic design reduces the risk of repetitive strain injuries, ensuring a more comfortable and efficient pipetting experience.
  3. Accuracy and Precision:
    • Micropipettes are renowned for their accuracy and precision in dispensing small liquid volumes. They are calibrated to deliver specific volumes within a specified range, ensuring reproducible results and reducing experimental variability. The precise measurement capabilities of micropipettes enable researchers to achieve accurate dilutions, sample transfers, and reaction setups, critical for obtaining reliable and meaningful data.
  4. Ease of Use:
    • Micropipettes are designed to be user-friendly and easy to operate. They typically feature simple and intuitive mechanisms for volume adjustment, aspiration, and dispensing. The pipette tips can be easily attached and ejected, allowing for quick and seamless transitions between samples. The user-friendly nature of micropipettes makes them accessible to both experienced researchers and novice users, facilitating efficient and error-free pipetting.
  5. Versatility:
    • Micropipettes offer a wide range of volume capacities, allowing researchers to handle various sample sizes and volumes. They can accurately dispense volumes ranging from microliters (µl) to milliliters (ml), making them versatile tools for a broad range of applications in molecular biology, microbiology, biochemistry, and other scientific disciplines. The availability of different pipette tip sizes and materials further enhances their versatility.
  6. Sample Preservation:
    • Micropipettes enable precise and controlled handling of delicate and valuable samples, such as DNA, proteins, enzymes, and drugs. The accurate measurement and gentle pipetting technique minimize sample loss, contamination, and damage, ensuring the integrity and preservation of valuable samples throughout the experimental process.

Overall, micropipettes provide significant advantages in terms of time savings, ergonomic design, accuracy, precision, ease of use, versatility, and sample preservation. These advantages contribute to improved efficiency, reliability, and reproducibility in laboratory experiments, making micropipettes indispensable tools for researchers and technicians in various scientific fields.

What you shouldn’t do with micropipettes?

  • You should not drop your pipette. If you drop your pipette, it could cause it to not be in the calibration. Be delicate and cautious.
  • Aspirate the contents into the pipette. It happens, we’ve seen it. If you’re working with chemicals that are harsh and it gets stuck in the pipette could cause to the build-up of substances and cause inaccurate results.
  • Move the dial up or below the volume limit. This could cause damage to the volume indicator as well as other parts of the pipette.
  • Jam pipette tip into pipette. This could not only harm the pipette’s tip but also more importantly, the pipette shaft. What this means is that you will most likely require a more fit pipette tip.

Make sure you treat your micropipette properly. It’s an extremely precise instrument, and, as such, careful care of its performance has to be observed. If you find that it’s out of performance or isn’t performing as it used to, it’s time to have it checked or calibrated. If you need further help regarding how to use a micropipette, we’ve created an extensive Guide to Proper Pipetting resources to better help you.

Precautions To Be Taken While Using Micropipettes

Micropipettes are pricey! To maintain their functionality, it is essential that they be handled with care. Please follow these rules to maintain their functionality:

  • Never alter the volume beyond the micropipette’s range. No micropipette should be set below zero microliters. Never adjust the P20 over 20  µ µL, the P200 above 200  µL, or the P1000 above 1 mL.
  • Never force the volume control dial to turn. If the knob gets difficult to adjust, you are likely surpassing the pipette’s capacity or the pipette is destroyed. Communicate the issue to your instructor.
  • Avoid dropping the micropipette.
  • Always use the micropipette with a steady motion. This will aid in providing accurate measurements and prevent pipette damage. There should not be any crackling sounds.
  • Always store pipettes vertically.
  • Never place a pipette containing liquid on the bench.
  • Always select the correct size pipette for the volume being measured.
  • Always dispose of cigarette ends in the correct waste receptacle.

Sources of errors in Micropipette

Errors in micropipette measurements can occur due to various factors. Here are some common sources of errors in micropipettes:

  1. Handling Error:
    • Inaccurate measurement can result from improper handling techniques. Inconsistent pressure applied to the plunger or improper attachment of the pipette tip can lead to variations in the dispensed volume. It is essential to maintain a steady and even pressure while operating the micropipette and ensure a secure attachment of the tip to minimize errors.
  2. Tip Quality:
    • The quality and compatibility of the pipette tips can significantly impact measurement accuracy. Tips that do not fit properly on the micropipette can cause leaks or improper sealing, resulting in inaccurate volumes. It is important to use high-quality tips that are designed specifically for the micropipette being used.
  3. Liquid Properties:
    • The properties of the liquid or sample being pipetted can introduce errors in measurement. Highly viscous liquids, for example, may cling to the walls of the tip and not be completely transferred, leading to under-dispensing. Certain liquids, such as volatile or corrosive substances, may interact with the pipette materials and affect their accuracy. It is crucial to select the appropriate type of pipette tips and ensure compatibility with the liquid properties.
  4. Calibration Errors:
    • Proper calibration of the micropipette is essential for accurate measurements. Calibration errors can arise from inadequate or incorrect calibration techniques. If the micropipette is not calibrated regularly or if the calibration process is not performed correctly, it can result in systematic errors in volume dispensing. Regular calibration using proper methods and equipment is necessary to maintain the accuracy of the micropipette.
  5. Environmental Factors:
    • Environmental conditions, such as temperature and humidity, can affect the performance of the micropipette. Changes in these factors can impact the viscosity and surface tension of the liquid, leading to variations in volume measurements. It is important to work in a controlled environment and consider the impact of environmental conditions on the accuracy of the micropipette.

To minimize errors in micropipette measurements, it is crucial to follow proper handling techniques, use high-quality and compatible pipette tips, consider the properties of the liquid being pipetted, regularly calibrate the micropipette, and maintain a suitable laboratory environment. By addressing these potential sources of errors, more accurate and reliable measurements can be obtained using micropipettes.

FAQs

What is a micropipette used for?

A micropipette is a laboratory tool used to measure and dispense small volumes of liquid. It is commonly used in a variety of applications, including molecular biology, biochemistry, genetics, and cell culture.
The micropipette consists of a tube with a calibrated volume scale and a tip. The user inserts the tip into the liquid they want to measure or dispense, and then uses the micropipette’s thumb wheel or electronic controls to draw up or release a specific volume of liquid. Depending on the design, micropipettes can measure and dispense volumes of liquid in the range of 0.1 microliters to several milliliters.
There are different types of micropipettes available to suit different applications, such as single-channel or multi-channel micropipettes for dispensing multiple samples at once, fixed-volume or adjustable-volume micropipettes, and manual or electronic micropipettes.
In addition, depending on the type of micropipette, it could be calibrated to measure volume with much more precision and accuracy, which is useful in experiments that require small liquid volume measurements like DNA/RNA isolation and quantitative PCR.

What is a micropipette?

A micropipette is a laboratory instrument used to accurately measure and transfer small volumes of liquid, typically ranging from microliters (µl) to milliliters (ml).

How does a micropipette work?

Micropipettes work based on the principles of air displacement. By adjusting the plunger, a specific volume of air is displaced, creating a vacuum that aspirates or dispenses liquid through a disposable pipette tip.

What are the common volume ranges of micropipettes?

Micropipettes are available in various volume ranges, such as 0.1-10 µl, 2-20 µl, 20-200 µl, and 100-1000 µl. The selection of the appropriate micropipette depends on the desired volume range for a specific application.

How do I select the correct micropipette for my application?

The selection of a micropipette depends on the required volume range and the accuracy needed. Consider the volume increments, accuracy specifications, and the compatibility of the micropipette with the desired pipette tips.

How do I calibrate a micropipette?

Micropipettes should be calibrated regularly to ensure accurate measurements. Calibration involves using a balance or gravimetric method to measure the weight of liquid dispensed by the micropipette and comparing it to the expected volume. Manufacturers typically provide guidelines for calibration.

Can I autoclave micropipettes?

No, micropipettes should not be autoclaved as the high heat and pressure can damage their delicate internal components. Only the removable parts, such as the pipette tips and adapters, can be autoclaved.

How should I handle micropipettes to maintain accuracy?

Handle micropipettes with care to avoid damaging their internal mechanisms. Avoid applying excessive force, dropping them, or exposing them to harsh chemicals. Always use appropriate pipette tips and ensure they are securely attached.

Can micropipettes be used with viscous liquids?

Micropipettes are generally designed for aqueous solutions. When working with highly viscous liquids, it is advisable to use micropipettes specifically designed for such applications or consult the manufacturer’s guidelines.

How can I prevent cross-contamination when using micropipettes?

To prevent cross-contamination, always use new and sterile pipette tips for each sample. Avoid touching the outside of the pipette tip, as this can transfer contaminants. Properly dispose of used tips and follow good laboratory practices for handling samples.

How often should micropipettes be serviced or calibrated?

Micropipettes should be serviced and calibrated regularly according to the manufacturer’s recommendations. It is essential to follow the maintenance schedule to ensure accurate and reliable performance.

References

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What is Karyotyping? What are the scope of Microbiology? What is DNA Library? What is Simple Staining? What is Negative Staining? What is Western Blot? What are Transgenic Plants? Breakthrough Discovery: Crystal Cells in Fruit Flies Key to Oxygen Transport What is Northern Blotting? What is Southern Blotting?
What is Karyotyping? What are the scope of Microbiology? What is DNA Library? What is Simple Staining? What is Negative Staining? What is Western Blot? What are Transgenic Plants? Breakthrough Discovery: Crystal Cells in Fruit Flies Key to Oxygen Transport What is Northern Blotting? What is Southern Blotting?
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