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Updated: Aug 9, 2025

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

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

Published on: February 16, 2014

SpyDisplay: A versatile phage display selection system using SpyTag/SpyCatcher technology.

Sarah-Jane Kellmann¹, Christian Hentrich¹, Mateusz Putyrski¹

¹Bio-Rad AbD Serotec GmbH, Neuried, Germany.

|February 21, 2023

Summary

SpyDisplay is a novel phage display method using SpyTag/SpyCatcher ligation for antibody fragment (Fab) display. This robust system enables efficient selection of high-affinity antibodies and facilitates new applications in protein display.

Keywords:

Fab antibody library SpyDisplay SpyTag/SpyCatcher antibody phage display

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Area of Science:

Biotechnology
Molecular Biology
Immunology

Background:

Phage display is a key technique for selecting proteins and antibodies from gene libraries.
Traditional methods rely on genetic fusion of displayed proteins to phage coat proteins.

Purpose of the Study:

To introduce SpyDisplay, a new phage display method utilizing SpyTag/SpyCatcher protein ligation.
To demonstrate the functional display and efficient selection of antibody fragments (Fabs) using SpyDisplay.
To explore novel applications of SpyDisplay beyond traditional phage display.

Main Methods:

SpyTagged antibody Fabs were ligated to filamentous phages displaying SpyCatcher fused to the pIII coat protein.
A library of antibody Fabs was cloned and expressed in engineered E. coli.
Phage panning was employed to isolate specific high-affinity antibody clones.

Main Results:

Functional and covalent display of SpyTagged Fabs on phage was confirmed.
High-affinity antibody clones were rapidly isolated using SpyDisplay and phage panning.
SpyTagged Fabs proved compatible with modular antibody assembly and direct assay testing.

Conclusions:

SpyDisplay offers a robust and versatile platform for antibody selection and protein display.
The method streamlines traditional phage display applications and enables new ones, including N-terminal display and display of TAT-exported proteins.