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Common Approaches for Centrifugation in the Laboratory

   |  February 15, 2023

Cell centrifugation is an indispensable step in common cell experiments. It not only allows adherent samples to settle completely at the bottom of the tube, but also helps confirm cell concentration. The centrifuged samples can be used for subsequent experiments such as primary cell culture, cell passage culture, single cell sequencing, etc.
Different approaches for centrifugal separation should be adopted according to the type of cells and organelles.

So, what are the common approaches for cell centrifugation?

01 Differential centrifugation
Differential centrifugation separates particles of different sizes and shapes in the heterogeneous mixture based on particles’ sedimentation rate while gradually increasing centrifugal force. It is typically used for laboratory sample separation, such as organelle mitochondria, chloroplasts, proteins and viruses.

Separation steps
Step 1: Set a relatively lower speed to allow high-density particles to settle to the bottom of the tube, while the medium-small sized particles in the mixture are still suspended in the supernatant.

Step 2: Collect the sediment. The resulting supernatant from step 1 could be centrifuged at a higher speed to separate relatively medium or small sized particles and so on.

Figure 1. Differential centrifugation

02 Density gradient centrifugation
Density gradient centrifugation, also known as zonal centrifugation, includes rate zonal centrifugation and isodensity centrifugation.

1.Rate zonal centrifugation
Rate zonal centrifugation is typically used to separate cells or organelles with similar density but different sizes. Load the density gradient media such as sucrose, glycerin, CsCl and Percoll into the centrifuge tube first. One thing worth noting is that the density of the sample particles must be higher than the density of any point in the gradient liquid column. Given the separated particles have different settling velocities in the gradient liquid, different components in the mixed sample can form their respective zones at different positions of the gradient liquid column, separation is therefore realized during centrifugation.

Figure 2. Rate zonal centrifugation

2.Isodensity centrifugation
Isodensity centrifugation is applied for separating particles with different densities. With sufficiently centrifugal force and time, cells or organelles can settle or float to the medium with the same density in a continuous gradient medium. The separation is achieved as cells stay and reach equilibrium.Percoll is widely adopted as the medium, while CsCl, Cs2SO4 and triiodoformyl-glucosamine can also achieve stable gradient after long-time centrifugation.

Figure 3. Isodensity centrifugation

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Figure 4. M1416R High-Speed Benchtop Refrigerated Centrifuge

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