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Luminex-based quantification of Alzheimer's disease neuropathologic change in formalin-fixed post-mortem human brain
C Dirk Keene1, Angela M Wilson1, Mitchell D Kilgore1
1Department of Pathology, University of Washington, Seattle, WA, 98104, USA.
Insights
New Luminex techniques quantify Alzheimer's disease (AD) biomarkers, amyloid beta and hyperphosphorylated Tau, in formalin-fixed paraffin-embedded (FFPE) tissues. This enables more precise correlation of neuropathology with clinical factors in AD research.
Area of Science:
- Neuropathology
- Biomarker Quantification
- Neurodegenerative Diseases
Background:
- Formally fixed, paraffin-embedded (FFPE) tissues are crucial for research but lack quantitative measures for neurodegenerative disease studies.
- Current methods like immunohistochemistry offer limited quantitative insights, hindering correlation with clinical factors such as age, genotype, and disease stage.
- Accurate quantification of neuropathological markers is essential for understanding Alzheimer's disease (AD) mechanisms.
Purpose of the Study:
- To develop and validate Luminex-based assays for quantifying amyloid beta and hyperphosphorylated Tau in FFPE brain sections.
- To enable highly quantitative and molecularly specific neuropathology measures in FFPE tissues.
- To overcome limitations in current FFPE tissue analysis for AD research.
Main Methods:
- Development of Luminex-based techniques for quantifying amyloid beta and hyperphosphorylated Tau.
- Validation of the assay using FFPE brain sections from 30 neuropathologically assessed AD cases (low, intermediate, high AD neuropathologic change).
- Demonstration of assay expansion to detect other targets like IBA1 and GFAP.
Main Results:
- The Luminex assay successfully quantified amyloid beta and hyperphosphorylated Tau in FFPE sections.
- High levels of amyloid beta were detected in the frontal cortex and striatum of high AD neuropathologic change cases.
- High levels of hyperphosphorylated Tau were detected in the frontal cortex and hippocampus of high AD neuropathologic change cases.
Conclusions:
- Luminex-based assays provide a novel, highly quantitative method for analyzing FFPE brain tissues.
- This approach supports multiplexed, molecularly specific neuropathology measures, advancing AD research.
- The validated assay can be expanded to detect diverse antigenic targets, facilitating deeper exploration of neurodegeneration mechanisms.
Abstract:
The vast majority of archived research and clinical pathological specimens are stored in the form of formalin fixed, paraffin-embedded (FFPE) tissues, but, unlike fresh frozen tissue samples, highly quantitative measures in FFPE tissues are limited to immunohistochemical and immunofluorescence thresholding image analysis studies, cell counting, and ordinal ranking systems. This poses a significant obstacle for clinical investigations that aim to correlate diagnostic markers of neurodegenerative diseases like Alzheimer's disease (AD) with parameters like age, gender, drug exposures, genotype, disease stage, co-morbidities, or environmental factors. To overcome this limitation, we have developed Luminex-based techniques and protocols for the quantification of amyloid β and hyperphosphorylated Tau in FFPE brain sections. We validated the Luminex assay in FFPE sections from prefrontal cortex, hippocampus, and neostriatum from 30 cases that underwent prior neuropathological diagnostic assessment of AD following the current NIA-AA recommendations for AD: 10 cases diagnosed as not or low, 10 cases as intermediate, and 10 cases as high AD neuropathologic change. Consistent with the neuropathologic assessment, Luminex assay detected high amounts of amyloid beta in the frontal cortex and striatum, and high amounts of hyperphosphorylated Tau in the frontal cortex and hippocampus, of cases with high AD neuropathologic change. This assay can be expanded to detect diverse antigenic targets of interest, as we show here with IBA1 and GFAP. This novel approach supports multiplexed highly quantitative, molecularly specific neuropathology measures to further explore mechanisms of neurodegeneration in AD.
