Epigenetic Haemogram for Precise Immune Cell Quantification
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Current methods for quantifying immune cells, such as flow cytometry and immunohistochemistry, face challenges including the need for fresh samples, imprecise protein marker thresholds, loss of surface markers in tissue cells, and inability to detect cells with intra- or extracellular markers, leading to imprecise and incomplete immune cell profiling.
Innovation Solution
An epigenetic haemogram method using a nucleic acid normalization standard with cell-type specific and unspecific markers, allowing for precise quantification of blood and immune cells independent of sample integrity and composition, through bisulfite conversion and qPCR analysis.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If flow cytometry or immunohistochemistry is used to quantify immune cells, then cell quantification can be performed, but the methods require fresh samples and lose surface markers in tissue cells
Solution Approach 1:
The patent replaces protein-based detection methods (flow cytometry, immunohistochemistry) with nucleic acid-based detection. By converting immune cell identification from protein marker detection to DNA methylation pattern analysis through bisulfite conversion and qPCR, the method eliminates requirements for fresh samples and intact surface markers, enabling reliable quantification from frozen or stored samples.
Solution Approach 2:
The patent changes the detection parameter from protein epitopes (surface markers) to epigenetic DNA methylation patterns. This parameter change allows detection of immune cells based on genomic DNA characteristics that remain stable in frozen and stored samples, rather than relying on protein markers that are lost during sample processing and storage.
2Adaptability or versatility
If protein markers are used for cell identification, then cell subpopulations can be detected, but imprecise thresholds lead to measurement errors
Solution Approach 1:
The patent substitutes protein marker-based identification with DNA methylation-based identification. By detecting methylation patterns at specific CpG sites through bisulfite conversion and qPCR, the method provides precise, binary detection (methylated vs. unmethylated) that eliminates threshold determination issues inherent in protein intensity-based flow cytometry.
Solution Approach 2:
The patent uses DNA methylation patterns as a stable copy or signature of immune cell identity. Instead of measuring variable protein expression levels that require threshold setting, the method detects fixed epigenetic marks that serve as reliable molecular copies of cell type information, enabling precise identification without arbitrary thresholds.
3Ease of operation
If surface markers are used for cell detection, then immune cells can be identified, but markers are lost in tissue cells during processing
Solution Approach 1:
The patent replaces surface protein marker detection with intracellular DNA methylation detection. By targeting epigenetic modifications in the nucleus rather than surface proteins, the method remains reliable throughout tissue processing, freezing, and storage, as DNA methylation patterns are preserved while surface markers are lost.
Solution Approach 2:
The patent uses DNA methylation patterns as an intermediary that preserves immune cell identity information throughout sample processing. These epigenetic marks serve as stable mediators that maintain cell type information from fresh through frozen and stored samples, unlike surface markers that are lost during processing.
4Productivity
If conventional blood count methods are used, then total white blood cell count can be determined, but differential counting is time-consuming and imprecise
Solution Approach 1:
The patent creates a universal DNA-based detection system that simultaneously provides both total cell count and differential cell count information. By using cell-type-specific DNA methylation markers in a qPCR assay, the method delivers comprehensive immune cell profiling (granulocytes, monocytes, B cells, T cells, NK cells) with the speed and precision of molecular detection, replacing both manual differential counting and flow cytometry.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
Provides a comprehensive and precise blood and immune cell count, overcoming assay performance differences and enabling accurate quantification of intact and non-intact cells, including those in frozen or stored samples, with clear genomic-based identification.
Implementation Method 1
detecting blood cells in a biological sample epigenetically
Implementation Method 2
quantifying said blood cells as detected using a normalization standard
Data Source
Figure 1
Figure 2
Figure 3
AI summary
The present invention provides an epigenetic haemogram, also referred to as an epigenetic blood cell count that identifies the quantitative, comprehensive picture of cellular composition in a biological sample, wherein advantageously a normalization standard is used. The normalization standard is a nucleic acid molecule comprising at least one marker-region being specific for each of the blood cells to be detected, and at least one control-region being cell-unspecific, wherein said regions are present in the same number of copies on said molecule and/or a natural blood cell sample of known composition. Furthermore, the present invention relates to a kit and the use of a kit for performing the epigenetic assessment of comprehensive, quantitative cellular composition of a biological sample. The biological sample is derived from e.g. a mammalian body fluid, including peripheral, capillary or venous blood samples or subfractions thereof, such as peripheral blood mononuclear cells or peripheral blood monocytes, or a tissue sample, organ sample, or from frozen, dried, embedded, stored or fresh body fluids or tissue samples.