Thrombus DNA Content Biomarker for Cardioembolic Stroke Identification
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Solution Overview
Problem
Current methods for determining the etiology of acute ischemic strokes, particularly in cases of large vessel occlusion, face inconsistencies due to the heterogeneity of thrombus composition and the limitations of semiquantitative histological analyses, which hinder the development of accurate diagnostic tools.
Innovation Solution
Quantitative biochemical assays are used to analyze the DNA and GPVI content in thrombus homogenates from patients, allowing for the differentiation between cardioembolic and non-cardioembolic strokes, with a focus on identifying cardioembolic strokes in patients with undetermined etiology using a DNA content threshold and DNA/GPVI ratio.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Ease of manufacture
If semiquantitative histological analyses using nonspecific staining methods are used to assess thrombus composition, then the analysis process is simpler and more accessible, but the measurement precision and reliability of etiology determination deteriorate due to inter- and intra-observer variability
Solution Approach 1:
The patent replaces manual histological staining and scoring methods with automated flow cytometry-based quantitative analysis. This substitution eliminates observer variability inherent in manual scoring while maintaining accessibility through standardized automated protocols. The flow cytometric measurement of thrombus components (RBCs, platelets, leukocytes, fibrin) provides objective, reproducible quantitative data that directly improves measurement precision without requiring complex manual intervention.
Solution Approach 2:
The patent transforms the analysis from semiquantitative histological scoring to quantitative biochemical parameter measurement. By changing the measurement approach to quantify specific thrombus component concentrations (RBC count, platelet count, leukocyte count, fibrin content) using flow cytometry, the system achieves higher measurement precision. These quantitative parameters serve as reliable biomarkers for etiology determination, resolving the contradiction between ease of analysis and measurement precision.
2Measurement precision
If quantitative biochemical assays are used to analyze thrombus composition, then the measurement precision and reliability of etiology determination improve, but the device complexity and analysis complexity increase
Solution Approach 1:
The patent employs a flow cytometer, which is a multi-functional device capable of performing multiple measurements simultaneously. The same instrument quantifies different thrombus components (RBCs, platelets, leukocytes, and fibrin) using various fluorescent markers and detection channels. This multi-functionality reduces the need for multiple specialized devices, thereby limiting the increase in device complexity while maintaining high measurement precision for comprehensive thrombus composition analysis.
Solution Approach 2:
The patent changes the measurement parameters from manual histological scoring to quantitative biochemical parameters measured by flow cytometry. By standardizing these parameters (RBC count, platelet count, leukocyte count, fibrin content) and using automated detection, the system achieves high measurement precision without proportionally increasing device complexity. The quantitative approach allows for objective, reproducible results that can be reliably used for etiology determination.
3Reliability
If thrombus composition is analyzed to determine AIS etiology, then diagnostic accuracy improves, but the heterogeneity of thrombus composition creates inconsistencies in diagnostic results
Solution Approach 1:
The patent addresses thrombus heterogeneity by changing from qualitative histological assessment to quantitative biochemical parameter measurement. By measuring specific concentrations of RBCs, platelets, leukocytes, and fibrin using flow cytometry, the system transforms the variable, heterogeneous thrombus composition into standardized quantitative data. These normalized parameters enable consistent diagnostic criteria that account for natural variations in thrombus composition, thereby improving diagnostic accuracy while managing the challenge of heterogeneity.
Solution Approach 2:
The patent employs a feedback mechanism where the quantitative thrombus composition data is systematically analyzed and compared against established diagnostic criteria. The flow cytometric measurements provide objective feedback that can be reliably interpreted to determine AIS etiology. This structured feedback approach ensures that despite thrombus heterogeneity, the diagnostic process remains consistent and reproducible, improving overall diagnostic accuracy through standardized interpretation protocols.
Data Source
AI summary
Determining acute ischemic stroke (AIS) etiology is crucial for guidance of secondary prevention. Here, the inventors performed a correlation analysis between AIS etiology and AIS thrombus cellular composition and content, as assessed using quantitative biochemical assays. In particular, homogenates of 250 AIS patient thrombi were prepared by mechanical grinding. Platelet, red blood cell, and leukocyte content of AIS thrombi were estimated by quantification of glycoprotein (GP)VI, heme, and DNA in thrombus homogenates. AIS etiology was defined as cardioembolic, non-cardioembolic, or embolic stroke of undetermined source (ESUS), according to the TOAST classification. Cardioembolic thrombi were richer in DNA (35.8 vs 13.8 ng/mg, p<0.001) and poorer in GPVI (0.104 vs 0.117 ng/mg, p=0.045) than non- cardioembolic ones. The area under the receiver operating characteristic curve of DNA content to discriminate cardioembolic thrombi from non-cardioembolic was 0.72 (95% Cl, 0.63 to 0.81). With a threshold of 44.7 ng DNA/mg thrombus, 47% of thrombi from undetermined etiology would be classified as cardioembolic with a specificity of 90%. In conclusion, thrombus DNA content may provide an accurate biomarker for identification of cardioembolic thrombi in AIS patients with ESUS.