Useful Notes on Freeze-Fractured Membranes (With Diagram)

The fluid-mosaic model of membrane structure is beautifully supported by visual evidence provided when freeze-fractured membranes are examined with the transmission electron microscope.

D. Branton, who pioneered this field, showed that membranes rap­idly frozen at the temperature of liquid nitrogen and cut or chipped with a microtome blade readily frac­ture along specific planes.

When the plane of the frac­ture intersects the plane of the membrane, the mem­brane is split along the center of the lipid bilayer, producing two “half-membranes” called the E half and the P half.

The E half is that portion of the mem­brane that faced the cell exterior, while the P half cor­responds to the portion that faced the “protoplasm” (i.e., cytosol).

One side of each half-membrane is the original membrane surface, called the E and P faces, whereas the other side is the newly exposed fracture face, called the E fracture face (EF) and P fracture face (PF). The fracture faces are extremely delicate and are not examined directly. Instead, a thin film of platinum and carbon is evaporated onto the surface of the fracture faces to produce a replica, which is then examined by transmission electron microscopy.

In many instances before the replica is made wa­ter (as well as other volatile materials) on or near the fracture surfaces is eliminated by sublimation (i.e., by carefully raising the temperature of the sample). This step, which used to be called “freeze-etching,” ex­poses additional surface features of the fracture face (see Fig. 15-9).

Electron micrographs of freeze-fractured cells show the membranes to be covered by numerous small par­ticles (Fig. 15-10). There is convincing evidence that the particles are membrane proteins (e.g., they disap­pear when the membranes are first treated with proteolytic enzymes).

This suggests that the plane of fracture passes around the protein molecules rather than through them. This relationship is depicted in Figure 15-11. The relatively uniform background ap­parent in the fracture face in Figure 15-10 corre­sponds to the surface of one-half of the lipid bilayer.