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Intracellular Amyloid-β Detection from Human Brain Sections Using a Microfluidic Immunoassay in Tandem with MALDI-MS
Jorvani Cruz Villarreal1,2, Keegan Kow1,2, Brian Pham1,2
1School of Molecular Sciences, Arizona State University, Tempe, Arizona 85287, United States.
Abstract:
Alzheimer's disease (AD) currently affects more than 30 million people worldwide. The lack of understanding of AD's physiopathology limits the development of therapeutic and diagnostic tools. Soluble amyloid-β peptide (Aβ) oligomers that appear as intermediates along the Aβ aggregation into plaques are considered among the main AD neurotoxic species. Although a wealth of data are available about Aβ from in vitro and animal models, there is little known about intracellular Aβ in human brain cells, mainly due to the lack of technology to assess the intracellular protein content. The elucidation of the Aβ species in specific brain cell subpopulations can provide insight into the role of Aβ in AD and the neurotoxic mechanism involved. Here, we report a microfluidic immunoassay for in situ mass spectrometry analysis of intracellular Aβ species from archived human brain tissue. This approach comprises the selective laser dissection of individual pyramidal cell bodies from tissues, their transfer to the microfluidic platform for sample processing on-chip, and mass spectrometric characterization. As a proof-of-principle, we demonstrate the detection of intracellular Aβ species from as few as 20 human brain cells.
Insights
Researchers developed a novel microfluidic immunoassay to detect intracellular amyloid-β (Aβ) species in human brain cells. This method allows for the analysis of Aβ in Alzheimer's disease (AD) research, offering new insights into neurotoxicity.
Area of Science:
- Neuroscience
- Biochemistry
- Analytical Chemistry
Background:
- Alzheimer's disease (AD) affects over 30 million people globally, with limited understanding of its pathology hindering therapeutic development.
- Soluble amyloid-β (Aβ) oligomers are implicated as key neurotoxic species in AD, but intracellular Aβ in human brain cells remains poorly understood due to technological limitations.
- Investigating intracellular Aβ in specific human brain cell types is crucial for understanding AD's neurotoxic mechanisms.
Purpose of the Study:
- To develop and validate a microfluidic immunoassay for the in situ mass spectrometry analysis of intracellular Aβ species.
- To enable the assessment of intracellular Aβ in archived human brain tissue at the single-cell level.
- To provide a novel tool for gaining insights into the role of intracellular Aβ in Alzheimer's disease pathogenesis.
Main Methods:
- Utilized selective laser capture microdissection to isolate individual pyramidal cell bodies from archived human brain tissue.
- Developed a microfluidic platform for on-chip sample processing of isolated cells.
- Performed mass spectrometry for the characterization of intracellular Aβ species within the microfluidic system.
Main Results:
- Successfully demonstrated the detection of intracellular Aβ species using the developed microfluidic immunoassay.
- Achieved detection of intracellular Aβ from as few as 20 human brain cells, showcasing high sensitivity.
- Validated the feasibility of analyzing intracellular Aβ directly from archived human brain tissue.
Conclusions:
- The novel microfluidic immunoassay enables sensitive detection and characterization of intracellular Aβ species in human brain cells.
- This technology provides a powerful new approach for studying the role of intracellular Aβ in Alzheimer's disease.
- The findings pave the way for deeper understanding of AD pathophysiology and the development of targeted diagnostics and therapeutics.
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