Epithelial Cell Quantification via Extended Antibody Incubation
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Solution Overview
Problem
Current methods for determining the concentration of epithelial cells in blood samples admixed with anticoagulants are inefficient, as they fail to account for the initial masked state of antigens specific for epithelial cells, leading to delayed accessibility and saturation in antibody binding.
Innovation Solution
A method involving erythrocyte lysis, suspension in a buffer, and prolonged incubation with labeled antibodies or antibody fragments directed against EpCAM and other epithelial cell antigens, allowing for increased accessibility and reliable determination of cell concentration over several hours.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional antibody binding methods are used with short incubation times, then the process is fast and efficient, but the antigen accessibility is limited due to initial masking, leading to inaccurate cell concentration determination
Solution Approach 1:
The patent applies preliminary action by performing erythrocyte lysis and antigen demasking treatments before the antibody binding step. This preliminary preparation removes masking substances from epithelial cell antigens, making them accessible for antibody binding. The method also includes a prolonged incubation period (at least 12 hours) to ensure complete antibody-antigen complex formation, thereby achieving accurate cell concentration determination without the time-loss problem of repeated measurements.
2Reliability
If prolonged incubation is implemented to achieve antigen saturation, then measurement accuracy improves, but the time required for the procedure increases significantly
Solution Approach 1:
The patent applies parameter changes by modifying the incubation time parameter to at least 12 hours and adjusting the temperature parameter (e.g., 37°C or 4°C) to optimize the antibody-antigen binding reaction. These parameter changes ensure complete saturation of antibody binding sites while maintaining reliable and reproducible results. The standardized parameter protocol allows the procedure to be performed reliably without requiring repeated measurements, thereby maintaining productivity.
3Measurement precision
If erythrocyte lysis and prolonged incubation are performed, then antigen accessibility increases and cell quantification becomes reliable, but the procedural complexity increases
Solution Approach 1:
The patent applies segmentation by dividing the detection procedure into distinct sequential steps: (1) erythrocyte lysis to remove red blood cells, (2) antigen demasking treatment to expose epithelial cell antigens, (3) prolonged incubation with labeled antibodies, and (4) quantification of epithelial cells. This segmented approach simplifies the overall procedure by making each step clear and manageable, reducing procedural complexity while ensuring accurate epithelial cell quantification.
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 enables accurate and reliable quantification of epithelial cells by extending the incubation time to achieve a decreasing binding rate, overcoming the initial masking of antigens and improving detection sensitivity.
Implementation Method 1
adding antibodies, antibody fragments or antibody mimetics which each bear at least one label and which are each directed against the epithelial cell adhesion molecule (EpCAM) and/or at least one other antigen specific for epithelial cells
Implementation Method 2
lysing the erythrocytes present in the blood sample or aspirate sample by addition of a buffer which brings about a lysis of the erythrocytes
Data Source
AI summary
A method for determining a concentration of epithelial cells in a blood sample or aspirate sample originating from a human being or mammal and mixed with anti-clotting agent. Here, following the addition of antibodies, antibody fragments, or antibody mimetica, which are each aimed against an antigen that is specific to epithelial cells, the sample is incubated until a decreasing binding rate of the binding of the antibodies, antibody fragments, or antibody mimetica to the cells is achieved. Only then is the number of marked cells and the original concentration of the cells in the blood sample or aspirate sample determined.