MICA-Binding Antibody Isolation via Fluorescent Cell Sorting
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Solution Overview
Problem
Current methods for obtaining antibodies with therapeutic potential from human subjects are limited by low frequency, slow proliferation rate, and low antibody secretion levels of B cells, particularly in cancer patients, necessitating the development of novel approaches for high sensitivity and efficiency in isolating these cells.
Innovation Solution
The development of therapeutic compositions comprising antibodies or antibody fragments that immunospecifically bind to MICA or angiopoietin, utilizing specific heavy and light chain variable region sequences, and peptides with conservative amino acid substitutions, which can be used to target and isolate immune cells from cancer patients, enhancing the identification and isolation of memory B cells.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional methods rely on in vitro culture of memory B cells to obtain antibodies, then antibodies can be measured in culture supernatants, but the method alters the memory B cell phenotype to resemble plasma cells with distinct functional properties
Solution Approach 1:
The patent uses fluorescently labeled antigens to pre-identify and isolate memory B cells before any culture expansion occurs. This preliminary identification step allows direct sorting of the target cells without requiring phenotypic changes during culture, thus maintaining the original memory B cell characteristics while enabling precise antibody measurement.
Solution Approach 2:
The patent extracts and isolates specific memory B cells using fluorescent-activated cell sorting (FACS) based on antigen binding. By taking out the specific memory B cells of interest from the mixed population before culture, the method avoids the phenotypic drift that occurs during extended culture periods while still obtaining sufficient material for antibody measurement.
2Quantity of substance
If in vitro culture is used to expand memory B cells, then antibody secretion levels increase, but the proliferation rate is slow and takes up to one week
Solution Approach 1:
The patent replaces the slow mechanical process of in vitro B cell culture and proliferation with a direct fluorescent labeling and sorting approach. By using fluorescent antigens to directly label and sort memory B cells, the method eliminates the need for slow cultural expansion while still obtaining sufficient cell numbers for antibody analysis.
Solution Approach 2:
The method performs preliminary identification and isolation of memory B cells using fluorescently labeled antigens before any expansion is needed. This allows the subsequent steps to proceed with already-isolated cells, eliminating the bottleneck of slow proliferation while maintaining adequate antibody secretion levels for measurement.
3Measurement precision
If fluorescent antigen-based methods are used to identify memory B cells, then isolation sensitivity improves, but the frequency of therapeutic antibodies remains low at one cell per million peripheral blood mononuclear cells
Solution Approach 1:
The patent uses fluorescently labeled antigens as intermediaries to bridge the gap between low-frequency therapeutic antibodies and detectable signals. The fluorescent label amplifies the detection capability, allowing even the rare one cell per million memory B cells to be identified and isolated with high sensitivity, thereby enabling recovery of therapeutic antibodies despite their low frequency.
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
These compositions enable the efficient capture and utilization of rare memory B cells, improving the isolation of antibodies with therapeutic potential, thereby enhancing cancer treatment options by targeting specific antigens like MICA and angiopoietin.
Implementation Method 1
an antibody or antibody fragment that immunospecifically binds to MHC class I polypeptide-related sequence A (MICA), or an epitope thereon
Data Source
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AI summary
The present disclosure provides, in part, compositions comprising peptides immunospecifically binds to defined binding partners, wherein the peptides comprise at least complementarity determining regions relating to the complementarity regions shown in Table 1.