Soluble oligomeric forms of amyloid-? (A?) have been isolated from brain, plasma and CSF and these assemblies appear to correlate with the severity of Alzheimer's disease (AD). To further understand how A?3 assembly state influences activity, we have developed structurally defined preparations of oligomeric and fibrillar A?31-42. In vitro, these preparations induce distinct neurotoxic, inflammatory, and synaptic plasticity responses. However, the relative contribution of fibrillar and oligomeric A? to the pathogenesis of AD remains unresolved. We hypothesize that A? exists in at least two states in the brain a diffusible oligomeric state, and an insoluble fibrillar (polymeric) state, and the ratio and clearance of these conformational species is altered in AD patients, promoting neurodegeneration. To dissect this mechanism, conformational-specific antibodies that can distinguish between fibrillar and oligomeric aggregation states of the peptide are critical. We used oligomeric A?31-42 as an antigen and an antigen/antibody screen has yielded one oligomer-specific monoclonal antibody (MOAB-1). MOAB-1 does not recognize fibrils by antigen/antibody blotting and ELISA. By Western analysis of SDS-PAGE, MOAB-1 does not recognize A? monomer in unaggregated or oligomeric samples, and little immunoreactivity is detected in the fibril samples. Accomplishing the following specific Aims will further test our hypotheses: ? 1. We propose to isolate and biochemically characterize additional monoclonal antibodies specific for oligomeric or fibrillar A?1-42. ? 2. We will determine the effect of selected antibodies on oligomer- or fibril-induced neurotoxicity, neuroinflammation, and neuroplasticity. We expect that oligomer-specific monoclonal antibodies will inhibit oligomer and not fibril-induced effects and vice versa for the fibril-specific antibodies. ? 3. We will test the specificity and suitability for immunohistochemistry (IHC) of selected antibodies in rat brains that have been stereotaxically injected with oligomeric or fibrillar A?1-42. We expect oligomeric antibodies to immunolabel only the brains injected with A? oligomers and not fibril-injected brains, and vice versa. ? 4. Selected antibodies will be utilized in quantitative IHC and Laser Scanning Confocal Microscopy to determine their localization in hippocampal sections from the brains of neuropsychologically well-characterized patients through the NU-CNADC Neuropathology Core at the Feinberg School of Medicine. ? 5. An ELISA capture assay will be designed, using existing and novel antibodies, to facilitate measurement of soluble A? oligomers in extracts of brain, CSF, and sera. ? ? ? ?
