Amyloid -protein (A) oligomers may be the proximate neurotoxins in Alzheimer's disease (AD). ''Oligomer'' is an ill-defined term because many kinds have been reported and they often exist in rapid equilibrium with monomers and higher-order assemblies. We report here results of studies in which specific oligomers have been stabilized structurally, fractionated in pure form, and then studied by using a combination of CD spectroscopy, Thioflavin T fluorescence, EM, atomic force microscopy (AFM), and neurotoxicity assays. A monomers were largely unstructured, but oligomers exhibited order-dependent increases in -sheet content. EM and AFM data suggest that dimerization and subsequent monomer addition are processes in which significant and asymmetric monomer conformational changes occur. Oligomer secondary structure and order correlated directly with fibril nucleation activity. Neurotoxic activity increased disproportionately (order dependence >1) with oligomer order. The structure-activity correlations reported here significantly extend our understanding of the conformational dynamics, structure, and relative toxicity of pure A oligomers of specific order.Alzheimer's disease ͉ toxicity A lzheimer's disease (AD) is the most common form of late-life dementia. Current estimates of AD incidence are Ͼ24 million worldwide, a number that is expected to double every 20 years, reaching 81 million in 2040 (1). AD is a slowly progressive disorder with insidious onset and progressive impairment of episodic memory and executive function coupled with aphasia, apraxia, and agnosia (1).The amyloid -protein (A) appears to play an essential role in the pathogenesis of AD. A is produced throughout life through posttranslational processing of the A precursor (APP). Familial forms of AD increase A production or the propensity of A to aggregate (2). The ''amyloid cascade hypothesis'' proposes that assemblies of A initiate a process leading to neuronal dysfunction and cell death (2). The most potent neurotoxic assemblies appear to be oligomeric, rather than fibrillar, in nature (3, 4). For example, oligomers extracted from AD brain potently impair synapse structure and function (5). The smallest of these oligomers appears to be dimeric (5). However, a systematic correlation of oligomer structure and neurotoxic potency has not been reported. This correlation is critical for the targeting and design of disease-modifying therapeutic agents.Efforts to establish rigorous structure-toxicity correlations have been hindered by the complex, dynamic equilibria displayed by A (for recent reviews, see refs. 6 and 7). To enable determination of the oligomer frequency distribution, we have used in situ chemical cross-linking to prevent oligomer dissociation or growth (8-10). Oligomers thus stabilized can be visualized and quantified by SDS/PAGE. However, in theory, the method also could be used to produce pure populations of oligomers of defined order † , enabling the biophysical and biological studies necessary to establish structure-toxic...