How To Use A Pipette?

How To Use A Pipette

Table of Contents

What is a pipette?

A pipette is a laboratory instrument that is used to draw up, measure, dispense, and transfer small and accurate volumes of liquid. Sometimes, pipettes that dispense volumes between 0.1 and 1000 μl (or more) are called micropipettes.

Pipettes are used by lab workers routinely on a day-to-day basis. It is essential to select the right instrument so that you work efficiently.

Pipette types

Let’s consider the types of pipettes. It is important to understand their differences in order to select the right product for your application.

Volumetric pipette

A volumetric pipette is a glass or plastic tube, typically between 1 and 100 milliliters. It offers a high level of accuracy.

The pipette is designed to deliver a fixed volume of liquids. The volume is marked with a horizontal line.

Volumetric pipette

As a rule, on a volumetric pipette, you will find some information, including how much liquid will be transferred if the liquid is drawn up to the calibration line on the neck of the pipette, the temperature at which the calibration was made, and if it is a TD or a TC pipette.

“TD” means that the pipette is calibrated to accurately deliver a specified volume of liquid.

“TC” denotes that your pipette is calibrated to contain a specified volume of liquid with no remainder.

Graduated pipette

A graduated pipette has volume values marked on the pipette wall in increments. With this pipette, it is easy to accurately measure and transfer a sample from one container to another.

There are two types of graduated pipettes: Mohr and Serological pipettes. They differ by the position of the first graduation mark. On a Serological pipette, the graduation marks start nearer the end of the tip.

Look at the picture below. A Serological pipette is located under a Mohr pipette.

Volumetric pipette

Pasteur pipette

A Pasteur pipette is used to transfer a small quantity of the liquid. Louis Pasteur, the French scientist, used these pipettes during his researches, hence the name. Some variants of this pipette were widely used to dispense eye drops into the eye. That’s why they are called eye droppers. Pasteur pipettes are sometimes referred to as transfer pipettes. Applications include lab use as well as dispensing small amounts of liquid medicines.

There are also other pipette types that are distinguished based on certain criteria.

Mechanical & electronic pipettes

Mechanical & electronic pipettes

Based on the operating mechanism, there are manually operated (mechanical) and electronic pipettes.

A mechanical pipette works based on the following principle. You have to press down the plunger to work with such pipettes. We will consider the working principle of a manually operated pipette in more detail a bit later (see the “How does a pipette work?” section). Mechanical pipettes are easy to be fully sterilized.

An electronic pipette is typically automated. You can dispense and aspirate liquids just pressing the button. Volume adjustment is done electronically. Such pipettes are more popular than their mechanical counterparts. Let’s find out why.

The benefits of electronic pipettes:

  • Fast and easy operation

Productivity and efficiency are crucial when working in the laboratory. With an electronic pipette, you will save your time for other important tasks.

  • Accuracy and precision are maintained

An electronic pipette will dispense the selected volume with a high level of accuracy and reproducibly as well.

  • Minimization of human error

The automation of the pipetting process is what you need for accurate results.

  • Ergonomics

Repetitive pipetting may cause musculoskeletal disorders. Electronic pipettes are perfect to minimize repetitive strain injury.

Fixed volume & adjustable volume pipettes

Fixed volume & adjustable volume pipettes

A fixed volume pipette allows you to work with a set volume of liquid, it is perfect if you perform one task repetitively and you don’t need to change the liquid volume. In addition, it is more accurate than an adjustable volume pipette (also known as a variable volume pipette), which is great for another purpose, e.g. if you have many tasks that require different volumes. Such pipettes have a special volume adjustment knob to easily set the volume you need.

Single-channel & multi-channel pipettes

Based on the number of pipette heads, there are single-channel and multi-channel pipettes. These pipettes can be either manually operated or electronic.

A single-channel pipette allows you to get accurate measurement results using one disposable tip.

Single-channel pipettes are perfect for laboratories with a low throughput of samples. If there are many samples to work with, consider using multi-channel options.

A multi-channel pipette is used to accurately measure and fill numerous vials of liquid at once. With these pipettes, you will increase your efficiency.

Electronic multi-channel pipettes are widely used in laboratories as identical liquid draws are achieved just pushing the button.

Positive displacement & air displacement pipettes

Based on the working principle, pipettes are classified into positive displacement and air displacement pipettes.

Take a look at the diagram below.

ositive displacement & air displacement pipettes

On the left, you can see an air displacement pipette, on the right - a positive displacement pipette.

An air displacement pipette has a small air cushion between the piston and the sample. These pipettes are suitable for standard applications, however, such factors as atmospheric pressure, temperature, density and viscosity of the liquid can affect the performance of your air displacement pipette.

A positive displacement pipette has a piston that is in direct contact with the sample. There is no air between the liquid and the piston. As a result, positive displacement pipettes are considered to be more accurate when it comes to volume measurement. Such pipettes are great for highly viscous or volatile samples.

What does a pipette consist of?

Let’s consider the elements of a fixed volume pipette. Look at the diagram below.

What does a pipette consist of

  • Plunger (or push button) is located on the top of the pipette and used to take up the liquid into the tip and dispense it from the tip.
  • Eject button (or tip ejector) is pressed to eject the tip.
  • Volume indicator (or volume display or digital readout) shows the volume of the liquid that is needed to be aspirated or dispensed.
  • Disposable tip is a replaceable part of the pipette. It is used to contain the liquid.

In addition to the above-mentioned elements, a variable volume pipette has a volume adjustment knob that is used to set the volume.

How does a pipette work?

Let’s consider the working principle of a pipette.

By using the plunger, the liquid can be taken up into and expelled from the pipette tip. The plunger has 2 stops.

It is pressed down to the 1st stop to aspirate the liquid up into the pipette tip. The tip of the pipette is immersed into the liquid with the plunger button being still pressed down. To draw the liquid up into the pipette, the plunger is slowly released.

When you press the plunger to the 1st stop, the internal piston displaces the air volume that is equal to the volume that you see on the volume display. When you release the pressure, it leads to the filling of liquid that is equal to the air volume removed as shows the volume indicator.

The plunger is pressed down to the 2nd stop to dispense the liquid that is in the tip.

The plunger is pressed down to the 2nd stop to dispense the liquid that is in the tip

When aspirating the liquid, there are two important factors that influence this process:

  1. the aspiration angle - it should be vertical, otherwise, too much liquid will be taken in;
  2. the immersion depth - the depth of the pipette tip inside the liquid - should be as small as possible, but at the same time, as large as necessary. If the pipette tip is immersed too deeply, additional liquid will be easily aspirated.

How to use a pipette?

How is a pipette used? Now that you know its working principle, it will be easy for you to understand how to use a pipette.

Things you need to know:

  • Do not point the pipette up as the liquid may run down into the pipette and, as a result, your pipette will be damaged and won’t function properly.
  • Release the plunger slowly when you withdraw the liquid. The liquid won’t rush into the end of your pipette and won’t clog it up.
  • Use the tip of the right size for each pipette.
  • Use a new tip for a new liquid.
  • Use the right pipette for the volume that is needed to be dispensed.

Things you will need:

Note that any piece of your personal protective equipment must be the right size and fit properly.

To use a pipette, follow the steps below:

1. Attach the tip to the pipette.

How to use a pipette

Open the box with the pipette tips, choose one, and apply light pressure to insert your pipette into the tip. Close the box with the pipette tips to avoid their contamination.

2. Press the plunger down to the 1st stop.

How to use a pipette

3. Immerse the tip into the liquid.

How to use a pipette

The tip of your pipette should be immersed into the liquid just slightly below the surface.

4. Check if the aspiration angle & immersion depth are correct (see the “How does a pipette work?” section). Then, slowly release the plunger.

How to use a pipette

5. Press the plunger down to the 1st stop again and aspirate the liquid once again.

How to use a pipette

Repeat this procedure 2-3 times in order to saturate the air in the tip.

When dispensing the liquid, make sure that the angle of your pipette is between 20 and 45 degrees. The tip should be in contact with the vessel wall.

6. Press the plunger down to the 1st stop.

How to use a pipette

7. Press the plunger further down to the 2nd stop to perform the blow-out.

How to use a pipette

Thus, you will empty the pipette tip completely.

8. Eject the pipette tip into the waste bin by pressing the tip ejector button on the pipette.

How to use a pipette

How to read a pipette?

Reading a pipette is easier than you think it is.

The range can be found on the body of the pipette or on its plunger. Among the most common ranges is 20 - 200 μl (microliters). In this case, the lowest volume you can work with is 20 μl and the highest volume for this pipette is 200 μl.

Note that it is not recommended to work with the volumes that are outside the indicated range as the springs will get worn out and as a result, your pipette won’t function properly.

As you know, a pipette has a volume indicator.

How to read a pipette

You can see the numbers but what do they mean? Let’s figure it out!

Note that we look at the pipette when its plunger is at the top and the pipette is not upside down.

In our example, we have 200 μl as the biggest value that can be read.

The highest number on the readout of this pipette is the hundreds place, the number on the readout below is the tens place, and the next number below is the ones place.

If you turn the volume adjustment knob, the lowest dial will change first, them the one above it and so on.

How to calibrate a pipette?

Things you will need:

The calibration of your pipette is important to ensure that you will get precise results.

Now, follow the steps below:

Part 1 - Check the calibration

1. Pour the distilled water into the beaker.

How to calibrate a pipette?

2. Measure the temperature of the water.

How to calibrate a pipette?

Immerse the thermometer into the water, leave it for at least 1 minute. Write down the temperature. Remove the thermometer and dry it off.

3. Put the weigh boat on the balance, zero it out.

How to calibrate a pipette?

You can use a balance with doors as there is an isolated chamber. In this case, you can put the weigh boat in that chamber and close the doors. If you have a regular balance with no doors, just put the weigh boat on your balance. Then, push the Zero or Tare button. Wait until you see zero on the display.

4. Prepare your pipette.

How to calibrate a pipette?

Wipe it down with ethanol, ensure nothing clogs the tip end. Attach the tip and set the volume for testing. It is recommended to test the smallest and the largest volumes. Remember that the range is indicated on the pipette.

5. Pre-rinse the tip prior to calibration.

How to calibrate a pipette?

Push the plunger down to the 1st stop and immerse the tip (about 0.08 in) into the distilled water. Release the plunger for liquid aspiration. Dispense the liquid by pressing the plunger. Repeat this procedure 3 times. Push the plunger down to the 2nd stop to dispense the remnants. Now the pipette can be removed from the water.

6. Draw up the calibration volume.

How to calibrate a pipette?

Make sure that the tip is not in the water. Press the plunger down to the 1st stop. Immerse the tip (about 0.08 in) into the distilled water. Release the plunger to aspirate the liquid. Wait for a second and then remove the tip from the water.

7. Dispense the liquid into the weigh boat on the balance.

How to calibrate a pipette?

The tip should be placed against the bottom of the weigh boat. Push the plunger down to the 1st stop. Move to the spot slightly away from the water and press the plunger down to the 2nd stop. Don’t release the plunger, lift the pipette tip away from the weigh boat. The tip should not contact neither any surface, nor your hands.

8. Write down the weight that is shown on the balance display.

How to calibrate a pipette?

Look at the balance display, wait until the numbers stop changing, and record the final result.

9. Repeat steps 1-8 at least 10 times.

How to calibrate a pipette?

When you have several results for the same volume, you can average them.

Part 2 - Calculate the results

1. Write down the formula for calculating the volume.

Calculate the results of pipette

Here’s the formula: V = w * Z

V - the calculated volume of the dispensed water;

w - the weight of the water;

Z - the conversion factor that is based on the water density.

2. Find out the conversion factor based on the water density.

Use Table 1.

Note that at sea level, standard air pressure in millibars is 1013.2.

Table 1. Values for Z (µg/mg), as a function of temperature and atmospheric pressure, for distilled water.

For instance, if the temperature of your distilled water is 20 degrees, then, the Z value will be 1.0029 µg/mg.

3. Average the results of weighing the water.

Calculate the results of pipette

The volume of the water dispensed by the pipette should be weighed at least 10 times. Add all the values you have (you should have 10) and divide by 10.

4. Plug the values into the formula.

Calculate the results of pipette

Use this formula: V = w * Z

5. Calculate the accuracy of your pipette.

Calculate the results of pipette

Use the following equation: A = 100 x Vavg/V0.

A - the accuracy of the pipette;

Vavg - the average calculated volume;

V0 - the value you set the pipette to dispense.

Note that the accuracy should be between 99-101%. If your pipette is calibrated the right way, the value you calculated should be close to the value that was set on the pipette. If the pipette didn’t pass the calibration test, contact the manufacturer of your pipette or a specialist that can calibrate your pipette.

How to maintain a pipette?

In order to keep your pipette in good condition, follow a few simple rules:

  1. Clean the pipette regularly. Otherwise, the dirt build-up can get in the way of your liquid handling operation.
  2. If you don’t really need to adjust the pipette, don’t do that, thus you will extend the lifetime of your pipette.
  3. Avoid reusing the tip of the pipette.
  4. Store the pipette in a clean and dry place.

Conclusion

Pipettes are widely used in laboratories. There are different pipette types for different applications. You need to make sure you have reliable pipettes that can offer high levels of accuracy and precision. Hopefully, now you know the elements of a pipette and understand how to use, read, calibrate, and maintain it.

Choose from an extensive range of top-quality pipettes on Mega Depot.

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