Related Experiment Video
Updated: Aug 18, 2025
Detecting Amyloid-β Accumulation via Immunofluorescent Staining in a Mouse Model of Alzheimer's Disease
Published on: April 19, 2021
Development of Alkaline Phosphatase-Fused Mouse Prion Protein and Its Application in Toxic Aβ Oligomer Detection
Kaori Tsukakoshi1, Rikako Kubo1, Kazunori Ikebukuro1
1Department of Biotechnology and Life Science, Graduate School of Engineering, Tokyo University of Agriculture and Technology, 2-24-16, Naka-cho, Koganei, Tokyo 184-8588, Japan.
Abstract:
Amyloid β (Aβ) oligomers play a key role in the progression of Alzheimer's disease (AD). Multiple forms of Aβ assemblies have been identified by in vitro and in vivo analyses; however, it is uncertain which oligomer is highly neurotoxic. Thus, understanding the pathogenesis of AD by detecting toxic Aβ oligomers is crucial. In this study, we report a fusion protein of cellular prion protein (PrPc) and alkaline phosphatase (ALP) from Escherichia coli as a sensing element for toxic Aβ oligomers. Since the N-terminus domain of PrPc (residue 23-111) derived from mice is known to bind to toxic Aβ oligomers in vitro, we genetically fused PrPc23-111 to ALP. The developed fusion protein, PrP-ALP, retained both the binding ability of PrPc and enzymatic activity of ALP. We showed that PrP-ALP strongly bound to high molecular weight (HMW) oligomers but showed little or no affinity toward monomers. The observation that PrP-ALP neutralized the toxic effect of Aβ oligomers indicated an interaction between PrP-ALP and toxic HMW oligomers. Based on ALP activity, we succeeded in detecting Aβ oligomers. PrP-ALP may serve as a powerful tool for detecting toxic Aβ oligomers that may be related to AD progression.
Insights
Researchers developed a novel fusion protein, PrP-ALP, to detect toxic amyloid-beta (Aβ) oligomers implicated in Alzheimer's disease (AD). This tool shows promise for identifying neurotoxic Aβ assemblies crucial for AD pathogenesis.
Area of Science:
- Neuroscience
- Biochemistry
- Molecular Biology
Background:
- Alzheimer's disease (AD) pathogenesis involves amyloid-beta (Aβ) oligomers, but the specific neurotoxic forms remain unclear.
- Detecting these toxic Aβ oligomers is critical for understanding AD progression and developing therapeutic strategies.
Purpose of the Study:
- To develop a novel biosensor for detecting toxic amyloid-beta (Aβ) oligomers.
- To investigate the potential of a fusion protein, PrP-ALP, as a tool for identifying neurotoxic Aβ assemblies.
Main Methods:
- A fusion protein (PrP-ALP) was engineered by combining the Aβ-binding domain of mouse cellular prion protein (PrPc) with alkaline phosphatase (ALP).
- The binding affinity of PrP-ALP to different forms of Aβ (monomers vs. high molecular weight oligomers) was assessed.
- The enzymatic activity of PrP-ALP was utilized to detect Aβ oligomers, and its neutralizing effect on Aβ-induced toxicity was evaluated.
Main Results:
- The fusion protein PrP-ALP demonstrated specific binding to high molecular weight (HMW) Aβ oligomers, with minimal affinity for monomers.
- PrP-ALP retained both the Aβ-binding capacity of PrPc and the enzymatic activity of ALP.
- The study confirmed that PrP-ALP could neutralize the neurotoxic effects of Aβ oligomers, suggesting a direct interaction.
- Detection of Aβ oligomers was successfully achieved by monitoring ALP activity.
Conclusions:
- The developed PrP-ALP fusion protein is a sensitive tool for detecting toxic HMW Aβ oligomers.
- PrP-ALP shows potential for advancing the diagnosis and understanding of Alzheimer's disease progression.
- This novel biosensor could aid in identifying specific Aβ assemblies relevant to AD pathogenesis.
