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Subcellular fractionation is used to obtain pure fractions of different cell organelles from a cell lysate.

Two commonly used methods  for subcellular fractionation are  differential centrifugation and density gradient centrifugation.

Differential centrifugation uses sequential centrifugation at progressively increasing speeds for size-based separation of organelles.

First, the sample is centrifuged at a low speed of around 400 to 600 x g to sediment large structures like nuclei and cell debris.

Then, the supernatant is spun at high speed ranging from 10,000 to 20,000 x g to pellet organelles such as mitochondria, lysosomes, and peroxisomes.

Further centrifugation cycles at speeds more than 80,000 x g can separate microsomes, membrane fractions, and even ribosomes.

However, differential centrifugation cannot separate organelles that are very close in size or density, such as mitochondria from peroxisomes.

In such cases, density gradient centrifugation, a more sensitive method, is a valuable tool.

This method  uses density gradients established using chemicals

such as sucrose or glycerol to separate organelles into distinct layers based on size, shape, or density within a single tube.