Page 1

Exercise 6. Cell Fractionation: Isolation of Mammalian Red Blood Cell Plasma Membrane Using Differential Centrifugation

 


A. Introduction

 

Image Source: Wellcome Images

The study of the structure, function and biochemistry of cellular compartments and organelles often requires the isolation and purification of these cellular structures. Cell fractionation is the process scientists use to produce fractions of functioning cellular components. The cell fractionation process involves two basic stages. First, the disruption of the tissue and gentle lysis of the cells, a process which creates a suspension of cell components, and second, the separation of the cellular components into purified fractions, a process usually accomplished through centrifugation.

For several reasons, mammalian red blood cell plasma membrane is the perfect cellular component on which to first learn basic fractionation techniques. One reason is that mature mammalian red blood cells are very easy to obtain. A second reason is that mammalian red blood cells are easy to lyse. And a third reason is that mammalian red blood cells have no internal membrane (erythrocytes have no nucleus and no organelles), and therefore there is no source of possible contamination of the plasma membrane fraction from other types of cellular membranes.

The scientific objective of today's laboratory activity is to produce a relatively pure fraction of mammalian red blood cell plasma membranes. Once purified, the membranes will be frozen for later analysis of their protein composition.