The lowering of natively analyzed Abeta42 in cerebrospinal fluid (CSF) is used as a diagnostic tool in Alzheimer's disease (AD). The presence of Abeta oligomers can interfere with such analyses causing underestimation of Abeta levels due to epitope masking. The aim was to investigate if the lowering of CSF Abeta42 seen in AD is caused by oligomerization.
Abeta42 was analyzed under both denaturing and non-denaturing conditions. An Abeta42 oligomer ratio was calculated from these quantifications. The presence of oligomers leads to Abeta42 epitope masking during non-denaturing assays, resulting in a higher ratio.
The Abeta42 oligomer ratio was used for the assessment of oligomerized Abeta in human CSF, after being evaluated in transgenic mouse brain homogenates. AD and mild cognitive impairment (MCI) samples displayed the expected decrease in natively measured Abeta42 compared to healthy controls and frontotemporal dementia, but not when analyzing under denaturing conditions. Accordingly, AD and MCI CSF had a higher Abeta42 oligomer ratio in CSF.
Combining denaturing and non-denaturing quantifications of Abeta42 into an oligomer ratio enables the assessment of Abeta oligomers in biological samples. The increased Abeta42 oligomer ratio for AD and MCI indicates the presence of oligomers in CSF and that the lowering of natively measured Abeta42 is caused by oligomerization.
"Unfortunately, Ab has a tendency to aggregate and, as mentioned earlier, these aggregates are considered to be crucial to the pathology. The capacity of immunoassays to properly detect these aggregates and accurately determine the total Ab concentration has been called into question  . ELISA measurements are based on the principle that each target protein will bind a single enzyme-conjugated antibody, resulting in a measurable signal that is proportional to the quantity of protein in the sample. "
[Show abstract][Hide abstract] ABSTRACT: According to the predominant theories, soluble amyloid-beta (Aβ) aggregates are the principal neurotoxic agents in Alzheimer’s disease pathology, making them a popular target for the development of therapeutics and diagnostic markers. One of the most commonly used methods for determining the concentration of Aβ is ELISA. However, ELISA was developed for monomeric proteins and may be ill-suited for detecting aggregates. Therefore, we investigated the effect of aggregation on the ELISA measurement and developed a novel chemical pre-treatment method, designed to disaggregate Aβ peptides, to improve the ELISA measurement of the total Aβ concentration.
"Attention is being directed towards the measurement and understanding of post-translational modifications of Aβ. Tg mice carrying the Arctic APP mutation provide an opportunity to study protofibrillar Aβ42; the development of protofibril-specific assays has permitted the quantification of CSF protofibrillar Aβ in these mice (Englund et al., 2007; Lord et al., 2009). Pyroglutamate Aβ (AβpE) may be an important pathogenic contributor to AD and a potentially useful biomarker . "
[Show abstract][Hide abstract] ABSTRACT: Alzheimer's disease (AD) represents an escalating global threat as life expectancy and disease prevalence continue to increase. There is a considerable need for earlier diagnoses to improve clinical outcomes. Fluid biomarkers measured from cerebrospinal fluid (CSF) and blood, or imaging biomarkers have considerable potential to assist in the diagnosis and management of AD. An additional important utility of biomarkers is in novel therapeutic development and clinical trials to assess efficacy and side effects of therapeutic interventions. Because many biomarkers are initially examined in animal models, the extent to which markers translate from animals to humans is an important issue. The current review highlights many existing and pipeline biomarker approaches, focusing on the degree of correspondence between AD patients and animal models. The review also highlights the need for greater translational correspondence between human and animal biomarkers.
Neurobiology of Disease 04/2013; 56. DOI:10.1016/j.nbd.2013.04.010 · 5.08 Impact Factor
"Though our conclusions are limited by the small number of subjects, our findings are consistent with the presence of an identifiable intermediate state between Aβ42 monomers and the deposition of Aβ42 in insoluble plaques during the presymptomatic phase of FAD. Our findings are consistent with the previous observation that at least a part of the decrement of CSF Aβ42 levels seen in AD patients is eliminated when Aβ42 levels are measured in denaturing conditions that decrease oligomerization . These results should be confirmed in larger groups of presymptomatic persons with FAD mutations as well as of those destined to develop late-onset AD using monoclonal conformation-dependent antibodies  and other techniques for measuring oligomers . "
[Show abstract][Hide abstract] ABSTRACT: Oligomerization of amyloid beta (Aβ) is a hypothesized step in the formation of plaques in Alzheimer's disease (AD) but has been difficult to demonstrate in vivo in humans. As persons destined to develop familial AD (FAD) due to fully penetrant autosomal dominant mutations are essentially certain to develop the disease, they provide the opportunity to identify oligomers during the presymptomatic stage of the disease.
We measured levels of Aβ(42) using a conventional immunoassay and prefibrillar, fibrillar, and annular protofibrillar oligomers using polyclonal conformation-dependent antibodies in the cerebrospinal fluid (CSF) of 7 persons at risk for inheriting FAD mutations. Levels of oligomers were compared between FAD mutation carriers and noncarriers.
Compared to 2 noncarriers, annular protofibrillar oligomers were elevated, prefibrillar and fibrillar oligomers trended towards elevation and Aβ(42) monomer trended towards being decreased in 5 FAD mutation carriers.
Our data provide evidence for an identifiable elevation of CSF oligomers during the presymptomatic phase of FAD.
Shawn Kile, William Au, Carol Parise, Kimberley Rose, Tammy Donnel, Andrea Hankins, Matthew Chan, Azad Ghassemi
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