Subcellular Fractionation

For study  metabolic and macromolecules processes  within  cells,  it  is frequently helpful to isolate one type of subcellular organelle from the rest  of  the  cell  contents   by  subcellular fractionation. Firstly, the plasma membrane (and if present, cell wall) has to be ruptured. The tissue or cell sample is suspended  in an isotonic sucrose solution  (0.25-0.32  M) buffered at the appropriate pH, to do this, and by homogenization  the cells are then broken open in a homogenizer or blender, by sonication,  or by subjecting them to high pressures (nitrogen bomb or French press). The first homogenization,  and the following  subcellular fractionation, are  carried  out  at 4°C  usually  in order  to minimize  enzymic  degradation  of the cell's constituents.  The sample of broken cells is frequently strained through muslin or other fine gauze to remove larger lumps of material before proceeding further.