Experiment #4: Cell Fractionation and Separation

You: ‘I believe that the centrifuge would be useful in an experiment that would involve cell fractionation and separation.’ 

Emma: ‘This was a bit general for me and that’s why it was not my first choice. Dr Smith can you please give an example?’

Dr Smith: ‘Imagine you have a liver tissue sample or a batch of cultured cells. The aim here is to break them open gently—usually by using a homogenizer—so that all the organelles and subcellular components spill out into a ‘tissue homogenate.’ At this point, the centrifuge becomes invaluable.’

Emma: ‘But would a single separation be sufficient? Cell components don’t all have the same density.’ 

Dr Smith: ‘You are right Emma and thanks for pointing this out! What we need to do is to spin the homogenate step by step at increasingly higher speeds. In this way, you can carefully isolate different parts of the cell based on their density. For instance, one spin might pellet the nuclei and more massive debris at the bottom of the tube, leaving lighter organelles and cytosolic proteins in the supernatant. Another, faster spin after collecting that supernatant could then separate mitochondria or other organelles. Eventually, you end up with layers (or ‘fractions’) that each contain different cellular structures.’

Differential centrifugation of cultured cells (https://biologynotesonline.com/differential-centrifugation/)

Did you consider all your options?