What are an ultracentrifuge and its applications?

An ultracentrifuge is a centrifuge that spins a sample at a very high speed (rpm), which can be used for preparative and analytical purposes. Since biomolecules exhibit random thermal motion, they are not affected by Earth's gravity (i.e. g). However, the isolated biomolecules showed significant sedimentation when they were subjected to high acceleration. Therefore, use an ultracentrifuge to accelerate the sample at 60,000 RPM and 200,000 x g to 150,000 RPM and 1,000,000 x g.

There are two types of ultracentrifuges: analytical ultracentrifuges and preparative ultracentrifuges.

As the name suggests, preparative ultracentrifuges are used to separate or precipitate biological particles, viruses, organelles, membranes, and biomolecules, among others. Analytical ultracentrifuges, on the other hand, are used for qualitative and quantitative samples with low sample loads.

Analytical ultracentrifuges use an optical system to measure concentration distributions correctly. It uses a xenon flash as a light source. As the rotor turns, an image of the cell is projected through an optical system onto a film or computer.

The concentration of the solution at various points in the cell is determined by absorbing light of the appropriate wavelength. This can be done by measuring the blackening of the film or by the deflection of the recorder of the scanning system and fed into a computer or analyzer. This will allow the operator to observe the separation of sample concentration (absorbance) from radial position (cm).

Two key experiments can be performed using an analytical ultracentrifuge: sedimentation velocity experiments, sedimentation equilibrium experiments

Settling velocity experiment:

The deposition velocity experiment was designed to explain the entire deposition process. This helps to understand protein size, shape, molecular weight, etc. It can also be used to study reversible chemical equilibria.

Settling Equilibrium Experiment:

These experiments were performed under conditions where the final homeostasis of the macromolecule is critical. Therefore, isopycnic density gradient centrifugation can be performed using an analytical centrifuge, where the particles will migrate to the region of the centrifuge tube with a density equal to that of the solution. Settling is balanced by diffusion of opposite concentration gradients, resulting in a time-independent concentration profile.

Preparative ultracentrifugation is used to separate molecules from samples or extracts. They are not used to analyze the sample, but to extract the desired protein or molecule from the sample on a large scale.

Preparative ultracentrifuges are commonly used to separate particles based on their density, separate or collect denser particles for collection in particles, and clarify particle-containing suspensions. Sometimes researchers can also use a preparative ultracentrifuge if they need the flexibility to change the type of rotor in the instrument.

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