Centrifugation Protocol

Introduction

A centrifuge is a common piece of laboratory equipment used to separate different components in a liquid sample. It achieves this by using centrifugal force, which is generated by spinning the sample at a high speed. Being able to separate solids from a liquid or separate liquids of different densities is essential to many different experimental procedures, such as minipreps and RNA extractions. Plus, centrifuges are useful for simply collecting liquid to the bottom of a tube. You will likely encounter several different types of centrifuges that can accommodate different sized containers, spin at different speeds, or keep samples at specific temperatures. This protocol will cover the general procedure and features to keep in mind when using any centrifuge. However, each centrifuge may have specific instructions so be sure to check in with someone familiar with your lab’s instrument before use.

Last Update: September 2022

Equipment

• Centrifuge
• Microfuge tubes or other suitable container
• Personal protective equipment (PPE) appropriate for the biosafety level of the lab space

Reagents

• Sample to be centrifuged
• Water

Background Information

Front view of an open tabletop centrifuge with labels for the core parts that would be found on almost any centrifuge.

Types and Parts

There are several types of centrifuges:

• Mini or Tabletop Centrifuges sit on your benchtop and typically hold the common 1.5 mL microfuge tubes used in labs, though some larger models can hold larger conical tubes. Depending on the centrifuge their speed ranges can vary, but are typically lower than some other centrifuge types.
• Floor Centrifuges sit on the floor and can be about bench height. They often spin at similar speeds to their smaller cousins, but can hold much larger containers.
• Ultracentrifuges are also generally floor sized centrifuges that can hold larger containers, but they can also rotate at much higher speeds and are used for more specialized techniques.

Regardless of the type of centrifuge you use, most will have essentially the same components. The photo to the left shows a front view of an open tabletop microcentrifuge with the lid open, the rotor with microfuge tubes in place, and the control panel where you can set the time and speed needed for your experiment.

Procedure

An example of a balanced tabletop centrifuge, with tubes evenly distributed around the rotor.

How to Centrifuge

Now that you know the different parts of a centrifuge, you’re ready to begin using the centrifuge.

1. Determine the type of centrifuge needed. This decision should be based on the size of tubes you are using and the speed at which you need to spin your samples. For example, spinning down samples in 1.5 mL microfuge tubes at 10,000 rpm would require a tabletop microcentrifuge like the one in the picture above.
2. Wear PPE appropriate for the lab space in which you are working. Even if your samples may not require specific PPE, you never know what else has been placed in the centrifuge before you and may still be on the equipment.
3. Before using the centrifuge for your samples, ensure that the centrifuge is clean and that everything appears to be working smoothly.
4. Place your sample tubes into the rotor so that they are distributed evenly around the center (see image to right). Even distribution ensures that the centrifuge is balanced. Using the centrifuge in an unbalanced state can damage the centrifuge and be dangerous for the user. If you do not have a balanced number of samples, you may need to add a blank. Blanks are often an extra tube filled with water to the same volume as your samples. If your samples are significantly higher density than water, then you may need to fill the blank based on weight instead of volume.
   Pro-Tip
   For microfuge tubes, place them with the lid tabs pointing up or out of the centrifuge so that the pellets form in approximately the same place in each sample.
5. Place the centrifuge’s internal lid on the rotor (if it has one) and then close the external lid.
6. Adjust the centrifuge settings according to speed and time (and temperature, if applicable) listed in your protocol.
   Pro-Tip
   Spin speed is often given as Relative Centrifugal Force (RCF) or Revolutions Per Minute (RPM), these are not the same units and are not directly interchangeable. If you need to convert between the two, use a conversion calculator or chart like the G Force calculator .
7. The centrifuge should come up to speed with a pleasant hum. If you hear loud noises or the centrifuge begins to wobble or shake, this could indicate an unbalanced or improperly loaded centrifuge. Immediately stop the centrifuge and investigate the issue.
8. When you are finished spinning your samples, carefully remove them from the centrifuge so as to not dislodge any pellets or re-mix liquids. In cases where you are separating solids from a liquid, pellets represent the solids in your sample while the liquid portion is known as the supernatant. Depending on your experiment, you may be required to save the pellet, the supernatant, or both.

Conclusion

Centrifuges are important tools for many different protocols. Improper use of a centrifuge can damage the equipment, your samples, or you, so be sure to keep your centrifuge clean and balanced.