The Specificity and Cross-Reactivity of Antisynaptosome Antibodies as Determined by Sequential Adsorption Analysis

Abstract

An improved iso-osmotic, Ficoll-sucrose, density-gradient technique for isolating synaptosomes, synaptic membranes, brain mitochondria and three forms of myelin from adult rat brain is described. Antibodies raised in rabbits to isolated synaptosomes, synaptic membranes, brain mitochondria and liver mitochondria, when assayed by means of sequential adsorption analysis, showed that a major class of determinants is shared by brain mitochondria and synaptic membranes, that another major class is shared by myelin and synaptic membranes and that a third class is shared by liver mitochondria and the various brain fractions. Besides these crossreactivities synaptosomes also displayed a class of distinctive determinants separate from those of myelin; brain mitochondria displayed a class of distinctive determinants separate from those of synaptic membranes and from liver mitochondria, and liver mitochondria displayed a class of distinctive determinants separate from those of brain mitochondria. Synaptosomes also combined with a small but definite amount of antibody in two different antisera that did not cross-react either with isolated synaptic membranes or brain mitochondria at the given concentrations, thereby revealing a distinctive class of synaptosome determinants, possibly conformational in nature.