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[An efficient and accurate method for counting target molecules in phage-display peptide library].

Ke-Geng Wang¹, Qing-Ren Zeng, Zheng-Yang Yu

¹Department of Parasitology, School of Medicine, University of South China, Hengyang 421001, China.

Zhongguo Ji Sheng Chong Xue Yu Ji Sheng Chong Bing Za Zhi = Chinese Journal of Parasitology & Parasitic Diseases

|October 6, 2011

Summary

Real-time PCR offers a faster and simpler method for determining bacteriophage titer compared to the double-layer agar plate (DLAP) method. This quick assay provides accurate phage quantification with reduced variability.

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A Protocol for Phage Display and Affinity Selection Using Recombinant Protein Baits

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Published on: February 16, 2014

Area of Science:

Microbiology
Molecular Biology
Virology

Context:

Accurate bacteriophage titer determination is crucial for various applications, including phage therapy and molecular biology research.
Traditional methods like the double-layer agar plate (DLAP) assay are time-consuming and can exhibit significant variability.
Developing rapid and reliable methods for phage quantification is essential for efficient research and application.

Purpose:

To compare the accuracy, efficiency, and variability of real-time PCR assay versus the double-layer agar plate (DLAP) method for determining bacteriophage titer.
To evaluate the suitability of both methods for quantifying low, intermediate, and high phage concentrations.

Summary:

Both DLAP and real-time PCR accurately measured bacteriophage titer across different concentrations.
The DLAP method required multiple replicates (10) with larger sample volumes (10 microliters) and showed higher within-assay coefficient variation (CV) ranging from 4.93% to 30.38%.
Real-time PCR required fewer replicates (3) with smaller sample volumes (1 microliter) and demonstrated significantly lower within-assay CV (0.02% to 0.25%), indicating superior precision and efficiency.

Impact:

Real-time PCR presents a more efficient, rapid, and precise alternative to DLAP for bacteriophage titer determination.
This advancement can accelerate research in phage biology, diagnostics, and therapeutic applications by enabling quicker and more reliable phage quantification.
The findings support the adoption of real-time PCR for routine bacteriophage quantification in laboratory settings.