Epigenetic Marker for CD4+ T-Lymphocyte Identification
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Solution Overview
Problem
Current methods face challenges in reliably detecting, identifying, and quantifying CD4+ T helper lymphocytes, particularly in routine applications, due to non-specific expression of CD4 protein on various cell types.
Innovation Solution
A method analyzing the bisulfite convertibility of specific CpG positions in the CD3+CD4+ T helper cell non-methylated bisulfite convertible region, with a convertibility of at least 90% indicative of CD4+ T-lymphocytes, using PCR and specific primer pairs to distinguish CD4+ cells from other cell types.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If CD4 protein expression is used to identify T helper lymphocytes, then detection can be performed, but non-specific expression on various cell types reduces reliability
Solution Approach 1:
The invention extracts and utilizes the specific epigenetic methylation pattern of the CD4 gene as a unique identifier for T helper lymphocytes, separating the identification marker from the non-specific CD4 protein expression that appears on multiple cell types. This methylation pattern is extracted as a cell-type-specific feature that reliably distinguishes T helper cells from other CD4-expressing cells.
Solution Approach 2:
The invention applies local quality by focusing on the specific epigenetic state (methylation pattern) of the CD4 gene locus rather than the general CD4 protein expression. This localized epigenetic marker provides cell-type-specific identification, transforming a non-specific protein marker into a specific cellular identifier through analysis of the local genomic region's methylation status.
2Productivity
If conventional detection methods are used, then identification can be performed, but the methods are not robust for routine applications
Solution Approach 1:
The invention replaces conventional mechanical/cellular detection methods (such as flow cytometry requiring cell purification and staining) with an epigenetic analysis approach. By substituting the detection mechanism to analyze DNA methylation patterns instead of protein expression on cell surfaces, the method achieves greater robustness for routine applications while maintaining high reliability.
3Measurement precision
If purification or staining procedures are required, then cell identification can be performed, but the process becomes more complex and time-consuming
Solution Approach 1:
The invention extracts the epigenetic methylation pattern information directly from genomic DNA, eliminating the need for cell purification and protein staining procedures. By taking out and analyzing the methylation status of the CD4 gene locus directly, the method achieves precise detection without the complex preparatory steps required by conventional approaches.
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
This method allows for precise and robust detection and quantification of CD4+ T-lymphocytes in blood and tissues, independent of purification or staining procedures, facilitating diagnostic applications in autoimmune diseases, cancer, and immunodeficiencies.
Implementation Method 1
analyzing the bisulfite convertibility of at least one CpG position in the CD3+CD4+ T helper cell specific non-methylated bisulfite convertible region
Data Source
AI summary
The present invention relates to a method, in particular an in vitro method, for identifying CD3CD4 positive T lymphocytes of a mammal, wherein the method comprises analyzing the bisulfite convertibility of at least one CpG position in the CD3+CD4+ T helper cell specific non-methylated bisulfite convertible region according to SEQ ID No. 1, wherein a bisulfite convertibility of at least one CpG position to at least 90%, preferably to at least 91% and more preferably to at least 92% and most preferred to at least 95% in the sample is indicative for a CD4+ T-lymphocyte cell, in particular a CD3+CD4+ T-lymphocyte cell. The present invention further relates to analyzing the bisulfite convertibility of at least one CpG position in the genes FLJ00060, FLJ38379, PPP6C, CD226, ZBTB7B and TNFAIP8 that are capable of positively identifying CD4 expressing cells in whole blood and segregate between CD4 and CD8 positive CD3 positive cells. Furthermore, the present invention relates to a kit for performing the above methods as well as respective uses thereof.