Enhancing propagation and purification efficiency of M13 bacteriophage for improved phage display applications

The global biosensors market is rapidly growing, driven by increasing demand for quick, affordable, and portable diagnostic tools across sectors, including healthcare, environmental monitoring, food safety, and biomedical research. The key to biosensor function is the biological recognition element (BRE), which determines the specificity, sensitivity, and reliability of the device. Traditional BREs, such as antibodies and enzymes, face significant limitations, including instability, high costs, and variability. Phage display technology offers a strong alternative, providing durable, stable, and highly specific peptides as BREs. However, it requires effective amplification and purification methods to produce high-quality peptide libraries. This study examines key factors affecting the amplification of filamentous M13 bacteriophage, highlighting the negative impact of high multiplicity of infection (MOI) caused by superinfection exclusion and reduced phage adsorption efficiency. Our findings indicate that the bacterial growth phase is the most important determinant of M13 amplification efficiency. Furthermore, post-infection PEG/NaCl precipitation followed by high-speed centrifugation significantly outperforms traditional filtration methods in purifying phages, maximizing recovery and viability. These findings present an optimized, reproducible, and scalable approach to M13 phage amplification, improving the effectiveness of phage display for developing advanced biorecognition elements. Ultimately, this research provides a foundational framework for more efficient biosensing and therapeutic applications, filling critical gaps in the current biosensor development landscape.