A Specific Enzyme-Linked Immunosorbent Assay for Measuring β-Amyloid Protein Oligomers in Human Plasma and Brain Tissue of Patients With Alzheimer Disease

Center for Neurologic Diseases, Brigham and Women's Hospital, Harvard Medical School, HIM 616, 77 Ave Louis Pasteur, Boston, MA 02115, USA.
Archives of neurology (Impact Factor: 7.01). 03/2009; 66(2):190-9. DOI: 10.1001/archneurol.2008.565
Source: PubMed

ABSTRACT To examine in vivo levels of beta-amyloid (Abeta) oligomers (oAbeta) vs monomeric Abeta in plasma and brain tissue of patients with sporadic and familial Alzheimer disease (AD) using a new enzyme-linked immunosorbent assay (ELISA) specific for oAbeta.
To establish the oAbeta ELISA, the same N-terminal Abeta antibody was used for antigen capture and detection. Plasma and postmortem brain tissue from patients with AD and control subjects were systematically analyzed by conventional monomeric Abeta and new oAbeta ELISAs.
We measured oAbeta species in plasma samples from 36 patients with clinically well-characterized AD and 10 control subjects. In addition, postmortem samples were obtained from brain autopsies of 9 patients with verified AD and 7 control subjects.
Oligomeric Abeta and 4 monomeric Abeta species in plasma samples from patients with AD and control subjects were measured by ELISA.
The specificity of the oAbeta ELISA was validated with a disulfide-crossed-linked, synthetic Abeta(1-40)Ser26Cys dimer that was specifically detected before but not after the dissociation of the dimers in beta-mercaptoethanol. Plasma assays showed that relative oAbeta levels were closely associated with relative Abeta(42) monomer levels across all of the subjects. Analysis of sequential plasma samples from a subset of the patients with AD, including a patient with AD caused by a presenilin mutation, revealed decreases in both oAbeta and Abeta(42) monomer levels over a 1- to 2-year period. In brain tissue from 9 patients with AD and 7 control subjects, both oAbeta and monomeric Abeta(42) levels were consistently higher in the AD cases.
An oAbeta-specific ELISA reveals a tight link between oAbeta and Abeta(42) monomer levels in plasma and brain. Both forms can decline over time in plasma, presumably reflecting their increasing insolubility in the brain.

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Available from: Dominic M Walsh, Jun 12, 2014
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    • "Instead, those studies showed that cognitive deficits were highly correlated with synapse loss [30] [31] [32], and suggested that an as-yet-unidentified toxin, but not amyloid plaques, was responsible for triggering memory loss in AD. Identification of soluble amyloid-b oligomers (AbOs) as synaptotoxins that accumulate in AD brains [33] [34] [35] [36] stimulated a paradigm shift in the field, with AbOs now considered the proximal toxins responsible for synapse dysfunction and memory failure in AD (for recent reviews, see [1,37–39]). In the following sections, we review recent findings linking the neurotoxic impact of AbOs and defects in brain insulin signaling in AD. "
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    Alzheimer's & dementia: the journal of the Alzheimer's Association 02/2014; 10(1):S26–S32. DOI:10.1016/j.jalz.2013.12.004 · 17.47 Impact Factor
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    • "However, due to the low concentration of A␤-oligomers, its detection is a complex task. Using different methods [7] [8] [9] [10] [11] [12] [13] [14] [15] [16] A␤-oligomers have been detected. By flow cytometry, we detected A␤-oligomers in CSF of non-demented patients [17]. "
    • "E-mail address: Assays with apparent selectivity for soluble oligomers over monomers and large insoluble assemblies of synthetic Ab have been reported, but most have not been rigorously validated on natural human oligomers in biologic samples [7] [8] [9] [10] [11] [12] [13] [14] [15]. Herein we describe and extensively validate two sandwich ELISAs with very high specificity for oligomeric Ab and sensitivity sufficient to allow detection of low concentrations of natural Ab oligomers in human brain samples. "
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    ABSTRACT: BACKGROUND: Soluble oligomers of amyloid ß-protein (Aß) have been increasingly linked to synaptic dysfunction, tau alteration, and neuritic dystrophy in Alzheimer's disease (AD) and mouse models. There is a great need for assays that quantify Aß oligomers with high specificity and sensitivity. METHODS: We designed and validated two oligomer-specific (o-) enzyme-linked immunoassays (ELISAs) using either an Aß aggregate-selective monoclonal for capture and a monoclonal to the free N-terminus for detection, or the latter antibody for both capture and detection. RESULTS: The o-ELISAs specifically quantified pure oligomers of synthetic Aß with sizes from dimers up to much larger assemblies and over a wide dynamic range of concentrations, whereas Aß monomers were undetectable. Natural Aß oligomers of similarly wide size and concentration ranges were measured in extracts of AD and control brains, revealing >1000-fold higher concentrations of Aß oligomers than monomers in the soluble fraction of AD cortex. The assays quantified the age-related rise in oligomers in hAPP transgenic mice. Unexpectedly, none of 90 human cerebrospinal fluid (CSF) samples gave a specific signal in either o-ELISA. CONCLUSIONS: These new o-ELISAs with rigorously confirmed specificity can quantify oligomer burden in human and mouse brains for diagnostic and mechanistic studies and for AD biomarker development. However, our data raise the likelihood that the hydrophobicity of Aß oligomers makes them very low in number or absent in aqueous CSF.
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