Antigen-Specific Plasma Cell Immortalization via Pre-Activation
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Solution Overview
Problem
Current methods for producing monoclonal antibodies, such as hybridoma technology and phage display, face limitations in generating diverse antigen-specific antibodies, with low yields and stability issues, and require extensive time and resources, while plasmacytoma technology is limited by clonal diversity and in vivo selection.
Innovation Solution
The method involves contacting lymphocytes with antigens to obtain immunized cells, selecting and activating them based on cell surface markers, and immortalizing them using activating agents or viral vectors to produce antigen-specific plasma cells and antibodies, which can be cultured in vitro or in vivo to increase the frequency and stability of antigen-specific clones.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If hybridoma technology is used to produce monoclonal antibodies, then antigen-specific antibodies can be generated, but the frequency of successful clones is extremely low (one per 10^6-10^8 starting cells) and the process requires extensive screening and subcloning
Solution Approach 1:
The patent applies preliminary action by pre-activating B cells with antigen and cytokines before fusion to generate hybridomas. This pre-activation enriches the splenocyte population for antigen-specific, activated B cells that are primed for fusion, thereby increasing the frequency of successful antigen-specific hybridoma clones and reducing subsequent screening time
Solution Approach 2:
The patent changes key parameters of the hybridoma generation process by modifying the state of B cells prior to fusion. Specifically, B cells are activated through antigen exposure and cytokine treatment (changing their functional state), and the fusion conditions are optimized to enhance hybridoma formation efficiency from activated cells
2Quantity of substance
If plasmacytoma technology is used to generate monoclonal antibodies, then antibody production can be achieved, but clonal diversity is limited by in vivo clonal selection and plasmacytoma development
Solution Approach 1:
The patent applies segmentation by dividing the plasmacytoma generation process into distinct stages: in vitro activation and enrichment of antigen-specific B cells, followed by in vivo plasmacytoma development. This segmentation allows preservation of clonal diversity during the in vitro activation phase before immune selection occurs in vivo
Solution Approach 2:
The patent uses an intermediary approach by introducing activated B cells into an immunodeficient host that lacks functional T cells and B cells. This intermediary system allows the activated B cells to develop into plasmacytomas without competing with or being selected against by the host's immune system, thereby preserving the diversity of the original activated cell population
3Reliability
If traditional hybridoma methods are used, then monoclonal antibodies can be produced, but the process requires months of in vitro culture to stabilize clones and ensure strong mAb production
Solution Approach 1:
The patent applies preliminary action by pre-activating B cells with antigen and cytokines before fusion, creating a population of activated B cells that are already primed for antibody production. This pre-activation reduces the time required for clone stabilization because the hybridomas derived from activated cells are more likely to be high-producing clones from the outset
4Ease of manufacture
If phage display with naïve libraries is used to generate monoclonal antibodies, then animal immunization is avoided, but antibody affinities are two to three orders of magnitude lower than traditional fusion technology
Solution Approach 1:
The patent applies preliminary action by pre-activating B cells with antigen and cytokines before fusion. This pre-activation enriches the splenocyte population for antigen-specific B cells that have already bound antigen with high affinity through natural selection in the immunized animal, thereby capturing high-affinity clones that would be rare or absent in naïve libraries
Data Source
AI summary
Disclosed are methods for producing immortalized antigen-specific plasma cells and antibodies which include depleting an immunized cell population of CD138-positive cells and activating the depleted cells. The methods may be used to improve the efficiency of obtaining immortalized antigen-specific plasma cells.