title: Preparation of fluorescently-labeled amyloid-beta peptide assemblies: the effect of fluorophore conjugation on structure and function.
abstract: Recent research has focused on soluble oligomeric assemblies of the 42 amino acid isoform of the amyloid-beta peptide (A beta 42) as the proximal cause of neuronal injury, synaptic loss, and the eventual dementia associated with Alzheimer's disease (AD). While neurotoxicity, neuroinflammation, and deficits in behavior and memory have all been attributed to oligomeric A beta 42, the specific roles for this assembly in the cellular neuropathology of AD remain poorly understood. In particular, lack of reliable and well-characterized forms of easily detectable A beta 42 oligomers has hindered study of the cellular trafficking of exogenous A beta 42 by neurons in vitro and in vivo. Therefore, the objective of this study is to fluorescently label soluble oligomeric A beta 42 without altering the structure or function of this assembly. Previous studies have demonstrated the advantages of using tapping mode atomic force microscopy (AFM) to characterize the structural assemblies formed by synthetic A beta 42 under specific solution conditions (e.g., oligomers, protofibrils, and fibrils). Here, we extend these methods to establish a strategy for fluorescent labeling of oligomeric A beta 42 assemblies that are structurally comparable to unlabeled oligomeric A beta 42. To compare function, we demonstrate that the uptake of labeled and unlabeled oligomeric A beta 42 by neurons in vitro is similar. AFM-characterized fluorophore-A beta 42 oligomers are an exciting new reagent for use in a variety of studies designed to elucidate critical cellular and molecular mechanisms underlying the functions of this A beta 42 assembly form in AD.
authors.affiliation: Department of Anatomy and Cell Biology, College of Medicine, University of Illinois at Chicago, Chicago, IL 60612, USA.
authors.forename: L M
authors.initials: LM
authors.lastname: Jungbauer
authors.forename: C
authors.initials: C
authors.lastname: Yu
authors.forename: K J
authors.initials: KJ
authors.lastname: Laxton
authors.forename: M J
authors.initials: MJ
authors.lastname: LaDu