Phage Display Library Selection Cycle Acceleration
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Solution Overview
Problem
Current phage display methods for selecting target-binding proteins are time-consuming and inefficient, producing excessive phage populations with only a small fraction being used in subsequent rounds, leading to prolonged cycles and high resource utilization.
Innovation Solution
A method involving forming phage-immobilized target complexes, separating non-binding phage, infecting host cells, and producing replicate phage in the presence of the target, allowing for rapid selection and amplification of target-binding proteins through multiple cycles with reduced phage input and shorter cycle times.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If traditional phage display methods are used with overnight growth and purification, then large quantities of phage (10^13 to 10^15) are produced, but the cycle time is prolonged to one to five days and resources are excessively utilized
Solution Approach 1:
The patent performs phage amplification in advance during the same day, completing the growth step before the next selection cycle begins. This preliminary action eliminates the need for overnight incubation and allows sequential cycles to be performed in rapid succession, reducing total cycle time while maintaining sufficient phage quantities for effective selection
Solution Approach 2:
The patent implements multiple selection cycles within a single day by using periodic, repeated rounds of binding, elution, and rapid amplification. Instead of performing one cycle every 1-5 days, the method enables 3-5 cycles per day by optimizing the timing and duration of each step, thereby accelerating the selection process without compromising selection quality
2Quantity of substance
If traditional phage display methods are used with overnight growth, then sufficient phage are produced for selection, but only 10^11 to 10^12 out of 10^13 to 10^15 produced are used, leading to high resource utilization and waste
Solution Approach 1:
The patent produces only the necessary amount of phage for each selection cycle rather than generating excessive quantities. By performing rapid amplification with controlled duration and using smaller culture volumes, the method produces just enough phage (10^11 to 10^12) for effective selection, eliminating the waste of producing 100-1000 times more phage than needed
Solution Approach 2:
The patent recovers and reuses the target compound after each selection cycle by eluting bound phage and regenerating the target for the next cycle. This recovery approach eliminates the need to discard and replace target molecules, reducing material waste and enabling multiple cycles with the same target pool
3Reliability
If multiple cycles of phage display are performed with traditional methods, then binding affinity is improved through selection, but the process requires one to five days per cycle, significantly prolonging the overall selection time
Solution Approach 1:
The patent maintains continuous selection pressure across multiple cycles by immediately proceeding from one cycle to the next without prolonged interruptions. The rapid amplification and streamlined protocols ensure that the selection process flows continuously, allowing 3-5 cycles to be completed in a single day while maintaining the cumulative selection pressure needed to enrich for high-affinity binders
Solution Approach 2:
The patent optimizes critical parameters such as amplification duration, phage-to-target ratio, and elution conditions to enable faster cycling. By adjusting these parameters, the method achieves sufficient phage amplification in hours rather than overnight, and maintains effective selection stringency despite the compressed timeline, thereby improving both speed and binding affinity
Data Source
AI summary
Disclosed are methods of selecting phage encoding a target binding protein. The methods can include forming a mixture comprising a plurality of diverse display phage, a target, and a support, and forming phage immobilized to the support, each of which comprises a phage which binds the target and the target immobilized to the support. Phage that do not bind to the target are separated. Host cells are contacted with the phage immobilized to the support via binding to the target so that the host cells are infected by the phage immobilized to the support. Replicate phage are produced from the infected cells in the presence of the target immobilized to the support, thereby forming replicate phage immobilized to the support via binding to the support. Replicate phage that do not bind to the target are separated. Host cells are contacted with the replicate phage immobilized to the support.