Human erythrocyte membranes break down in a specific way when exposed to a solution of 1% potassium phosphotungstate (KPT), pH 6.5, as commonly used for negative staining. This process was examined as follows: KPT-treated ghosts were centrifuged to give a pellet and a supernatant solution. The latter was fractionated on a column of Sepharose 4B, using KPT as solvent, into three protein-containing fractions (I, II, III). The ghosts, pellet, supernatant, and fractions I, II, and III were analyzed biochemically (for protein, phospholipid, and sugar content, and for polypeptide and amino acid composition), and being in KPT, were examined directly by negative staining electron microscopy. It was found that KPT solubilizes the membrane proteins corresponding to bands 1, 2, 4.2, 5, and 6 on gel electrophoresis. Bands 1 and 2 (spectrin) are isolated in fraction II and bands 4.2, 5, and 6 are found in fraction III. Band 3, the PAS-staining bands, and phospholipid remain in the pellet. In its effect on the membrane KPT is similar to protein perturbants and to changes in the ionic environment. Electron microscopy reveals spectrin as long, straight fibers of diameter about 2 nm. Fraction III contains globular particles ranging in diameter from 5.5 to 11.0 nm. The pellet is composed of vesicles. Other minor structural elements are described. An ordered network structure was observed in the ghost preparation. It is composed of oriented fibers, similar to spectrin, that are cross-linked. The significance of this ordered network structure is discussed. A possible mechanism for the action of KPT is suggested.